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MACiE Version History

The first digit represents major database content changes, the second number represents minor edits to the database content, the final number represents changes to the website. Note version changes are listed with the current version first


Version 3.0

Version 3.0.11

This version (August 2011) represents the addition of 55 new entries, taking the total number of entries in MACiE to 335, and the incorporation of dataset comparison views, please see the FAQ and publication (in preparation) for more details and 3D information on the active site residues. It is also now possible to search for catalytic dyads and triads from within the amino acid residue searches. Links to the IUBMB ExplorENZ database and MetaCyc have now been added. Due to ProCognate no longer being updated, we have removed the link to it from the entry overview page. We have updated the documentation pages to fix some minor spelling mistakes and to reflect the updated structure of MACiE in the MySQL version.

Some specific changes have also been made to existing entries:

  • M0003. Overall comment on the formation of the active site added. Quinone has been re-annotated as a cofactor which is not regenerated in this reaction.
  • M0003. Overall comment on the regeneration of reduced glutathione has been added. Glutathione has been re-annotated as a cofactor.
  • M0013. Corrected the reaction step comments to be more descriptive.
  • M0017. Overall comment added relating to the different types of PNP enzymes.
  • M0019. Overall comment updated to include the fact that the configuration of the anomeric carbon is inverted during the course of this reaction.
  • M0032. The glutathione has been re-annotated as a cofactor. This mechanism only represents one half of the reaction to detoxify methyl-glyoxylate. A second enzyme uses the lactoylgluthathione product to produce D-lactate and regenerate glutathione.
  • M0048. Reference PMID:11258886 has been added, along with an overall comment stating that "It has been suggested [3] that a general base is not required, but a strong hydrogen bond with the N7 of the purine substrate provides sufficient transition-state stabilisation to permit relatively efficient catalysis". Further, it has been proven that Lys165 (previously annotated as a charge stabiliser) is not essential for catalysis, and is involved in substrate binding only (PMID:9521733) thus it has been removed from the list of catalytic residues. Finally, there is no mention of Arg169 in any of the literature references cited, this is likely to be a typographical error, and the residue should be Arg199, which is hydrogen bonding to the phosphate group. However, this residue is not mentioned in any of the other literature citations, and is so likely to be an error, thus the residue has been removed from the annotation.
  • M0082. In step 1 there was a stutter (duplicated deprotonates) which has now been removed. Also, the overall catalytic residues were incorrectly annotated, with Glu281 being listed as Gly281, which has now been corrected.
  • M0089. Phe76 was missing in the overall catalytic residue annotation due to the fact that it was incorrectly annotated as Phe66, this has now been corrected.
  • M0090. Step 1, Cys194 was incorrectly annotated as a hydrogen bond donor. This has been corrected to a polar/non-polar interaction and a comment has been added to the step.
  • M0091. The KEGG compound ID C00442 has changed to C00620.
  • M0098. Step 2 had an incorrect identifier for the cofactor (OF1_2002 instead of OF1_2001), this has been corrected.
  • M0108. The KEGG compound ID C03815 has changed to C00658.
  • M0110. The side chain of Ser1335 should not be charge stabilising, this is the function of the main chain amide in step 1.
  • M0117. Cys630 in step 1 should be a transition state stabiliser rather than an activator as the comment clearly states that it is stabilising the transient charge associated with the hydride transfer.
  • M0119. The aromatic thiamine ring rearrangement was shown in a different conformation to all the other thiamine diphosphate dependent reactions in MACiE, and whilst both rearrangements are equally valid (thus it's not a mistake per sae) the rearrangement has been modified to bring it in line with the other thiamine diphosphate MACiE entries. Further, no evidence in the references for the Thiamine diphosphate isomer in steps 3 to 7 where the methyl group on the five-membered ring is connected by a double bond, rather than single, and so this has been removed from the annotation. The Mg(II) cofactor which is associated with the thiamine diphosphate cofactor was missing and has now been included. Asn996 has also been added as an electrostatic stabiliser for the pyruvate and activated thiamine diphosphate moieties, to bring the entry in line with the CSA (as PDB:2pda).
  • M0126. A discrepency between the cofactor identifiers in the overall reaction and reaction steps has been corrected.
  • M0133. In steps 1, 2 and 3 the putidareoxin was incorrectly drawn with the number noting the number of negative charges as substcript, rather than superscript. This has now been corrected.
  • M0135. The KEGG compound ID C00425 has changed to C05422.
  • M0136. Step 2, O2- atom in description corrected to read 2- ion. Step 3, the spelling mistake in the description has (sinle should be single) been corrected.
  • M0157. The glutathione component in the substrate has been re-annotated as a cofactor. This mechanism represents the second step in a two step detoxification of methyl-glyoxylate, where glutathione was initially employed to sequender methyl-glyoxylate. In this reaction the glutathione cofactor is regenerated on hydrolysis. M0158. Two catalytic residues, Lys123 and Ser150 have been included in the annotation.
  • M0159. The KEGG compound ID C01891 has changed to X00137.
  • M0183. The DNA product has now been given an X number, rather than the generic code for DNA, to ensure that the reaction balances.
  • M0192. In step 1, Cys113 lowers the pKa of Cys110 by donating a hydrogen bond, so it has now been given the function of increase acidity (activator) (ref: 15854652).
  • M0203. The structure for X00117 was incorrect, this has now been corrected. With thanks to Christoph Mayer for identifying this bug.
  • M0207. The catalytic residue His455 was incorrectly annotated as His445 in the overall reaction and first two steps.
  • M0211. The stage images showed two main chain residues which had subsequently been found not to be catalytic and so were removed. Lys14 and Lys74 were also in the wrong protonation state (neutral where they should have been positive) in the images. The unidentified base in steps 2 and 5 has been identified as a hydroxonium ion.
  • M0212. A discrepency between the cofactor identifiers in the overall reaction and reaction steps has been corrected.
  • M0214. The catalytic residue Asn101 was incorrectly identified as Asn102 in steps 1 to 4.
  • M0217. Ser70 was incorrectly annotated as Ser80 in the overall reaction and Thr11 was annotated as Th11.
  • M0222. Tyr363 was incorrectly annotated in all the steps as Tyr36.
  • M0223. A discrepency between the cofactor identifiers in the overall reaction and reaction steps has been corrected.
  • M0233. The EC number has been corrected to change the 0's for hyphens.
  • M0234. The stereochemistry of the glutamine substrate and glutamic acid product have been corrected to L- enantiomer.
  • M0240. The substrate was incorrectly names protein lysine, when it should be protein acetyllysine. This has now been corrected. The EC number has been corrected to change the 0 for a hyphen.
  • M0241. Tyr82 incorrectly annotated in step 1 as Tyr75. In step 2, only one the two portions of the Cys1 residue was annotated.
  • M0248. The small organic acid has been re-classified as a substrate, rather than a cofactor.
  • M0249. Step 4 was annotated with PLP285 rather than PLP450, this has now been corrected.
  • M0250. The cofactor had been given an incorrect PDB location assignment, this has been corrected.
  • M0252. Lys53 was incorrectly annotated in the overall reaction as Lys51.
  • M0253. The catalytic lysine (Lys214) was missing from the stage images, this has now been corrected.
  • M0264. Error in bond order changes in step two corrected. A proton has been added to the overall reaction to ensure charge and mass balance.
  • M0274. Error in bond order changes in steps one and two corrected. The chemical structure of thiamine diphosphate was wrong in the underlying RXN files, which has now been corrected.
  • M0275. Error in bond order changes in steps one, two and eleven corrected. Steps 11 and 12 were annotated with the cofactor HET code TPP rather than TDP, this has now been corrected.
  • M0276. Cys140 was missing in the overall annotation. The PDB file has been changed from 2nap to 1ogy as this is the only PDB code that contains both structural units.
  • M0278. The image for step 1 showed the product copper in the incorrect oxidation state, this has now been corrected. The comments for step 1 and step 2 were identical. This was an error and has now been corrected.
  • M0279. The overall substrate and product were incorrectly drawn (the adenine was connected through the six membered ring rather than the five membered ring. This has now been corrected. Tyr209A was incorrectly annotated as Tyr204A in the overall reaction. Cys239B was incorrectly annotated as Cys558B in step 4.
  • M0280. The thiamine diphosphate was drawn in an incorrect conformation, this has been corrected, and as a consequence the bond order changes in steps one and three have also been corrected.

Version 2.5

Version 2.5.10

This version (December 2010) represents the addition of 20 new entries, taking the total number of entries in MACiE to 280. This version also includes some minor updates to existing entries and links to the CoFactor Database where appropriate.

Some specific changes have also been made to existing entries:

  • M0002. Added reference PMID:10716727, which suggests thatthe protonation ofthe beta-lactam nitrogen is the first step of the reaction and also added a comment to that effect in step 2, where the protonation currently occurs.
  • M0004. Changed the "R" groups which were representing the pseudoazurin to Cu+ and Cu2+ for the reduced and oxidised forms respectively. Changes the molecule id for oxidised pseudoazurin to the PDB code: 1bqk and added the reference for the pseudoazurin crystal structures (PMID:10364229). Fixed the stage reaction files to ensure that the copper ions involved in the reaction are in the correct oxidation states.
  • M0005. The protonation states of the overall reaction were incorrect. This has now been fixed.
  • M0009. Step 1 - corrected the mechanism image to show the His14 acting as the base. The comment has also been amended to make it clear that the cleavage of the C5'-O5' bond and formation of the C5'-S bond along with the function of the His14 residue occurs in a concerted manner.
  • M0011. Added an inferred return step in order to regenerate the zinc activated water molecule.
  • M0020. The acceptor has been identified as the tetraheme Cytochrome CymA in reference PMID:12899636 (with thanks to Graeme Reid for pointing this out). This reference has also been added. The electron acceptor in the substrates and products were also the wrong way around. This has been fixed.
  • M0028. The substrates and products are incorrect, should have three phosphate groups attached to the sugar ring, these have been corrected in the overall compound gifs and the stage reactions.
  • M0029, step 3 had incorrect bond change annotation. This has been fixed.
  • M0031 - Asp221 is essential for orientating the cofactor correctly, as well as stabilising the conversion of intermediate II to intermediate III (step 5). Trp82 has been included as a catalytic residue as it is involved in stabilising the reactive intermediate/transition states in the hydride transfer steps (reference 5 in MACiE). References PMID:2223755 and PMID:12525150 have also been added, along with comments on the final hydride transfer step and the conversion of intermediate II to III.
  • M0034. Step 5 had incorrect manual bond changes. This has been fixed.
  • M0039. Step 2 (the inferred return step) was incorrectly drawn. The assumption was made that the His241 and Asp14 were too far away from one another for a direct proton transfer, thus two waters were assumed to perform this role, which had been missed out of the image. This has now been corrected. Step 1, the manual annotation of teh bond changes were incorrect, this has now been corrected.
  • M0054. Step 7, the manual annotation of the bond changes were incorrect, this has been corrected.
  • M0055. Step 3, the manual annotation of the bond changes were incorrect, this has been corrected.
  • M0059. Steps 1, 3, 4 and 5, the manual annotation of the bond changes were incorrect, this has been corrected.
  • M0065. The overall reaction was unbalanced at the proton level. This has now been fixed. Further, Lys16 has been added as a catalytic residue. Evidence suggests that, whilst not essential, it may be involved in stabilising the intermediates of the first half-reaction (the phosphoryl transfer) as it is hydrogen bonded to the gamma-phosphate of the GDP substrate. We have also added reference PMID:9115248.
  • M0066. Steps 2 and 11, the manual annotation of the bond changes were incorrect, this has been corrected.
  • M0067. The overall reaction was unbalanced at the proton level. This has now been fixed. Further, the manual annotation of the bond changes in Step 4 were incorrect, this has been corrected.
  • M0069 - Added reference PMID:17698807, which implicates the main chain amide of Gly143 in stabilising the oxyanion hole formed during the course of this reaction, correspondingly, Gly143 has been added to the catalytic residues.
  • M0071. Changed the MACiE reaction for an alternative mechanism for which there is more evidence. Added reference PMID:10946228.
  • M0072. The overall reaction is reversible, not unknown. This has been corrected.
  • M0073. The compound identifiers for the substrate (X00026) and product (X00027) where updated for their KEGG identifiers C15522 and C00268 respectively. Step 04 had incorrect manual annotation of the bond changes, this has been corrected
  • M0082. The overall reaction was unbalanced at the proton level. This has now been fixed.
  • M0086. The overall reaction was unbalanced at the proton level. This has now been fixed.
  • M0087. Details of the alternative mechanisms proposed by Benini et al. and Barios et al. for this enzyme have been added (thanks to Dr Ciurli for pointing out that we had missed the alternative mechanism proposed by his group). The following references have also been added: PMID:20471401, PMID:15038715, PMID:11713685, PMID:10368287, PMID:16584179, PMID:17676790 and PMID:11300826.
  • M0088. Cys160 has been shown to not be required for catalysis, however, Ser161 (both the main chain amide and side chain portions of the residue) has been shown to be essential. Thus, Cys160 has been replaced by Ser161 and reference PMID:10820011 has been added.
  • M0089. Added a proton to the overall products, to correctly balance the reaction as shown in the steps.
  • M0090. Step 1, the manual annotation of the bond changes were incorrect, this has been corrected.
  • M0092. The overall reaction was unbalanced at the proton level. This has now been fixed.
  • M0093. Added a proton to the overall products, to correctly balance the reaction as shown in the steps.
  • M0099. Two steps have been added to the mechanism in which the PQQ cofactor is re-generated ready for another round of catalysis. A new reference has been added (PMID:12686102). Also, PQQ has been added as a cofactor (rather than a substrate). Further, a suggested alternative mechanism has been identified (PMID:15273299) and details of this have been added to the comments and PMID:15273299 has been added as a reference.
  • M0102. Steps 5 and 6, Tyr254 has been labelled as a hydrogen bond donor (to FMN). This annotation was missing.
  • M0103. Step 5, this step was incorrectly annotated as a proton relay. This has been corrected.
  • M0110. Step 1 had incorrect manual bond changes. This has been fixed.
  • M0112. Step 2 had incorrect manual bond changes. This has been fixed.
  • M0113. The protons and charge did not balance in step 6. This has been corrected.
  • M0116. Added a comment on teh conformational change of the reducing equivalents and reference PMID:12791694.
  • M0118. Step 1 had incorrect manual bond changes. This has been fixed.
  • M0119, step 5 was missing the radical formation annotation in the mechanism. This has been corrected.
  • M0120. Step 2 had incorrect manual bond changes. This has been fixed.
  • M0124. Steps 2, 5, 8 and 10 had incorrect manual bond changes due to long proton relay chains. This has been fixed.
  • M0127. Steps 1 and 2 had incorrect manual bond changes due to long proton relay chains. This has been fixed.
  • M0131. Steps 2 and 7 had incorrect manual bond changes. Step 2 due to long proton relay chains, step 7 was missing a single O-H bond formed. This has been fixed.
  • M0136. Step 4 had too many O-H bonds formed/cleaved in the manual annotation. This has been fixed.
  • M0138. Step 2 had incorrect manual bond changes due to long proton relay chains. This has been fixed.
  • M0141. Steps 1, 3, 6, 8 and 11 had incorrect manual bond changes due to long proton relay chains. This has been fixed.
  • M0143. Step 2 was missing the As-O bond order change in the manual annotation. This has been fixed.
  • M0148. The product (D-sedoheptulose 7-phosphate) was missing a CH2 group in the overall reaction. This has now been corrected.
  • M0150. Added stabilising and activating residues Glu122, Tyr56, Lys16 and Asn119. Added references PMID: 10353838 and PMID:10200157
  • M0156. Step 2 had incorrect manual bond changes. This has been fixed.
  • M0159. The reaction for step 1 was chemically unbalanced, this has now been corrected.
  • M0161. The 3'-hydroxy DNA product was missing the CH2 group between the sugar and the hydroxy group. This has now been corrected.
  • M0162. The product (cyclobutane pyrimidine dimer 5'-phosphate) was in an incorrect protonation state. This has now been corrected.
  • M0183. Step 2. The reaction description was incorrect: the MTHF doesn't donate an electron, but activation energy to the FAD. This has been corrected.
  • M0190. Tyr104 was missing from the overall list of catalytic residues, this has now been corrected.
  • M0204. Step 2 was missing one of the bond order changes in the manual annotation. The reaction for step 4 was missing the product water. This has now been corrected.
  • M0205. His377 was missing its side chain activity. This has been fixed.
  • M0208. The step reaction images were incorrect, these have now been updated to reflect the mechainsm as annotated. Steps 3 and 5 had incorrect manual bond changes due to a proton relay chain. This has been fixed. Step 5 also had the proton donor annotation missing for His202.
  • M0212. The images for the mechanism did not match the annotation. This has been corrected.
  • M0215. Step 1 a N-H bond was incorrectly annotated as a O-H bond change in the manual annotation. Step 4, the bond order change in the Asp28 was missing in the manual annotation. This has been fixed.
  • M0217. Step 1, the manual annotation of the bond formed/cleaved changes were incorrect, this has been corrected.
  • M0225. In step 2 of the manual bond annotation, the C-N bond formed should be a C-N bond cleaved. This has been fixed.
  • M0226. In steps 1 and 4, one of the C-C bond order changes in the manual annotation should be a C-N bond order change. This has been fixed.
  • M0228. The reaction for step 3 was unbalanced, further in step 3 one of the C-C bond order changes in the manual annotation should be a C-N bond order change. This has now been corrected. The manual annotation of the bond order changes in step 1 were incorrect, this has been corrected.
  • M0234. The pyrophosphate product in step 1 was in the incorrect protonation state. In step 4, one of the O-H bonds formed was missing in the manual annotation. This has now been corrected.
  • M0237. In step 1, one of the N-H bonds formed was missing in the manual annotation. This has been fixed.
  • M0239. In steps 2 and 4, the Fe-O bond order change missing in the manual annotation. This has been fixed.
  • M0243. Step 6 was chemically unbalanced, this was corrected (a methyl group was missing from the product intermediate. The substrates in steps 7 and 8 have also been corrected (as the mistake from step 6 was proagated through). Step 6 had incorrect manual bond changes due to proton relay chain.
  • M0245. Steps 2 and 5 had incorrect manual bond changes. Step 2 was missing the bond change to H. Step 5 was missing one of the O-H bonds cleaved. This has been fixed
  • M0247. The protonation state of the product in step 6 was incorrect. Steps 3, 6 and 8 had incorrect manual bond changes. Step 3 was missing an N-H bond formed in the manual bond changes. Step 6 was missing the C-S bond cleaved. Step 8 was missing the N-H bond formed. This has now been corrected.
  • M0249. Steps 2 and 11 had incorrect manual bond changes. Step 2 had extra N-H bonds formed/cleaved in the manual bond annotation. Step 11 had a missing N-H bond formed. This has been fixed.
  • M0252. The reaction for step 5 was missing the product water and hydroxonuim ion. Step 2 was missing some of the bond order changes in the manual annotation. This has now been corrected.
  • M0253. Added reference PMID:8986756, which suggests that Lys214 is important in stabilising the reactive intermediates and is suggested to be deprotonated (neutral). Added Lys214 to the catalytic residues.
  • M0255. Step 1, one of the C=C bond order changes should be C=N, also missing some of the bond changes due to a proton relay chain. This has been fixed.
  • M0256. The protonation state of the His51 in the product state of step 1 was incorrect. Step 1, one of the C=C bond order changes should be C=N, also missing some of the bond changes due to a proton relay chain. This has now been corrected.
  • M0260. Step 14 had incorrect manual bond changes. This has been fixed.

Version 2.4

Version 2.4.10

This version (September 2009) represents the addition of 37 new entries to the database. We have added titles into the references shown and added brought the references between Metal-MACiE and MACiE into agreement. We have continued to add descriptions of the steps to MACiE with all steps now having a description. We have also added GO terms based on the PDB code used and the associated UniProt codes. All overall reactions are also annotated with a reversibility tag, which depends on whether one or more steps are annotated as being either unknown (takes precedence over the other tags), irreversible (takes precedence over reversible reactions), or reversible. Finally, we have added a catalytic metal overview for MACiE entries with a corresponding Metal-MACiE entry. We have also included a version history for an individual MACiE entry, as well as this overview of all changes. A link to PROCOGNATE has also been added to the top level page of a MACiE entry. Finally, this version represents the re-formatting of the web pages to the EBI format.

The following changes have been made to MACiE queries: We have added in a query in order to identify the reaction steps in MACiE that have been assigned as the rate determining steps. This query is available from the Mechanisms query. We have also updated the Cofactor query, it is now possible to search for a cofactor performing a given function (and to specify whether one is interested in metal, non-metal or either type of cofactor). The PDB code search has been taken out of the index.pl search page and moved to a stand-alone search. This can be accessed directly at: http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/MACiE_NEW/overallQuery/pdb.pl?pdb=11as and will run exactly as before with only one addition: it will also check to see if the homologue identified in the CSA is associated with the same SwissProt identifier as the MACiE entry. The advancedEC search has also been moved to: http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/MACiE_NEW/overallQuery/advancedEC.pl?ec1=3&ec2=1&ec3=1&ec4=1. The EC number search has also been extracted from the index.pl script and can now be run from: http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/MACiE_NEW/overallQuery/ec.pl?ec1=3&ec2=1&ec3=1&ec4=1. This search has the exact functionality of the search when it was part of the index.pl script, i.e. in order to search for an EC sub-subclass, just input values for the first three variables. The amino acid query now incorporates a search that identifies which amino acid residues (and their MACiE entries) which act through both their side chain and main chain portions.

More specific changes include MACiE compound X00013 renamed as C01432, MACiE compound X00017 renamed as C00718, MACiE compound X00027 renamed as C00268, MACiE compound X00030 renamed as C00647, MACiE compound X00040 renamed as C00161, MACiE compound X00052 renamed as C00030, MACiE compound X00097 renamed as C15972, MACiE compound X00098 renamed as C16255 and the standardisation of the cofactor name thiamin diphosphate to thiamine diphosphate.

Some specific changes have also been made to existing entries:

  • M0002. Oxyanion hole stabilising residues (the main chain amide of Ser70 and the main chain amide of Ala237) have been added to the stage images.
  • M0006. PDB code has been changes from 3grs to 2gh5, which is the same enzyme (Homo sapiens), but contains both of the catalytic chains, and is the only crystal structure to do so.
  • M0014. His496 is now correctly annotated as a spectator residue, step 6 was incorrectly annotated as a coordination reaction, this has now been corrected to a substitution reaction. We have also added a new reference (PMID:15964077) which states that the step which produces the hyphalous acid is actually the rate determining step (step 6 in the MACiE entry). We have also included the halogenarion of an organic substrate in the entry to bring the overall reaction into line with the EC code as well as the other chloride peroxidase entries in MACiE.
  • M0016. Asp90 in step 4 is now correctly annotated as a spectator residue.
  • M0017. KEGG code C00442 has been replaced by C00620 within KEGG, this has not been reflected in MACiE.
  • M0019. KEGG code C05876 has been deprecated within KEGG, and replaced by G00011, this has been reflected in MACiE. Also for this reaction, the "alcohol" product (C00069) becomes G10694.
  • M0034. Step 7 was incorrectly annotated as a heterolysis. This has now been corrected.
  • M0039. The substrate (and product) of the reaction were incorrect and have now been fixed.
  • M0059. In the original citation for the entry (PMID:9685163) Arg260 was identified as the base for step four and the acid for step five. However, there is no Arg260, but Arg259 and Glu260. In the original entry for MACiE Arg259 was used as the base/acid. However, a later paper (PMID:12614613) and examination of the crystal structure suggest that Glu260 is the base. The original entry was therefore edited to reflect this.
  • M0062. Step 6 was incorrectly annotated as a heterolysis. This has now been corrected.
  • M0063. Step 6 was incorrectly annotated as a heterolysis. This has now been corrected.
  • M0068. The mechanism for step 3 appeared to be unsupported by the literature. Whilst we can find little direct evidence for the exact mechanism of this step, by analogy to other single electron reductions of FAD in MACiE, we have assumed that the deprotonation of the FAD facilitates the second single electron transfer from FAD to the ETF.
  • M0073. The substrate was incorrect (missing an NH2 group, this has now been corrected.
  • M0078. Ser244 was incorrectly annotated according to reference 3, it's function has been changed from steric to electrostatic stabiliser (it's role is to stabilise the His235 in it's active conformation such that he His235 is able to stabilise the reaction's transition state).
  • M0082. Steps 7 and 8 had incorrect annotation, this has now been fixed
  • M0105. The annotation for Glu869 was corrected in step 3.
  • M0106. In order to bring this thiamine diphosphate reaction into line with the other thiamine diphosphate reactions steps 1 and 2 (cofactor activation) have been combined into a single step, as have steps 8 and 9 (cofactor regeneration).
  • M0108. Steps 2 and 3 have been re-assigned as reversible.
  • M0110. The correct amino acid substrate has been corrected to the correct stereoisomer, i.e. the D-amino acid.
  • M0111. Steps 9 and 10 have been re-assigned as reversible.
  • M0122. The EC number 1.8.4.6 has been obsoleted and the entry transferred to EC 1.8.4.11.
  • M0123. Residues Trp234, Asn74 (side chain) and Arg265 were all incorrectly annotated as electrostatic stabilisers, when they are in fact acting to increase the electrophilicity of the sulfur atom.
  • M0126. The mechanism for [NiFe] Hydrogenase was incorrect (with thanks to Prof. Richard Cammack for this bug report). Two new references have been added: PMID: 17715982 and PMID: 18633545 which describe the more correct mechanism. There is still some discussion as to which metal ion the hydrogen molecule binds to initially. The residues Glu18B, Cys65B, Cys533B, Cys68B, Ser486B and His72B have been added. Glu18B is thought to be involved in proton shuttling, the other residues are all important in stabilising and activating the metal centre. Whilst we known that Cytochrome c3 is the physiological electron acceptor for this particular hydrogenase, it is a tetraheme cytochrome with all the hemes having different potentials, and probably only one or two are likely to accept electrons from the enzyme under physiological conditions, thus the electron acceptor has been renamed to a generic acceptor, rather than a specific one.
  • M0127. The electron acceptor/donors were incorrectly identified as ferrocytochrome c3 and ferricytochrome c3. These have now been renames to reduced ferredoxin and oxidised ferredoxin. An overall comment was added to the effect that the MACiE direction is the opposite to that cited by the EC number, but that the reaction is bi-directional and will take either 2H+ + 2e- → H2 OR H2 → 2H+ + 2e- Other notes: This type of hydrogenase may utilise either ferri- ferro cytochrome or ferredoxin as the acceptor. The dithiolate ligand is also more likely to be aza-dithiolate (S-CH2-NH-CH2-S) and the iron-sulfur clusters have been labelled with their correct PDB HET groups. It has also been explicitly noted that the proton relay chains in steps 1 and 2 is only tentatively identified and have been greyed out in the images.
  • M0132. Step 7 was incorrectly annotated as returning the enzyme to its native state. However, the mechanism utilises an unidentified base, so this cannot be certain and thus has been removed.
  • M0144. Step 5 was incorrectly annotated as returning the enzyme to its native state. However, the mechanism utilises an unidentified base, so this cannot be certain and thus has been removed.
  • M0168. His152 in step 2 is an electrostatic stabiliser, this annotation has now been correctly added.
  • M0187. An inferred return step has been added.
  • M0198. The acyl-CoA product was one carbon shorter than the long-chain carboxylate substrate, this has now been corrected.
  • M0213. PDB code changed from 1xfc to 1l6g to ensure that all the catalytic residues are present in the PDB file, and also to bring into line with the majority of the references.
  • M0219. Glu418 was incorrectly annotated in steps 5 and 6, this has now been corrected. A new reference Esakova et al. "Effects of transketolase cofactors on its conformation and stability", PMID:16125202 has been added, which states that the native metal cation is in fact Ca(II) rather than Mg(II). The overall comment and stage diagrams have been modified to reflect this change.

Version 2.3

Version 2.3.9

List of entries for version 2.3

This version (July 2008) represents the addition of 10 new entries and the incorporation of non-metal cofactor annotation. We have removed the static CML files, and now automatically generate the CML from the relational database. Currently, these files only include the textual annotation in the CMLReact format, i.e. do not include the chemistry. However, the chemistry will be added soon.

We have also started to include textual descriptions of the reaction steps on the overall reaction page. Currently this has only been completed for the ligases (EC 6) and entries 132 to 223.

We have also added to the current homologue listings (taken directly from the CSA) and now incorporate homologues based off the PDB and SwissProt, also allowing for more (or less) strict definitions of the homologues. This has been done with FastA and the help of Bill Pearson.

Navigation buttons have been added to the step view. These allow the user to cycle through the steps in an iterative manner using next and previous steps.

Some specific changes have also been made to existing entries:

  • M0020. The stage reaction mechanism images have been corrected.
  • M0062. The reaction arrows for stage 4 were incorrect, these have now been fixed.
  • M0134. 4a-hydroxytetrahydrobiopterin now has a KEGG identifier (C15522)
  • M0136. Added reference 3, PMID:8331655
  • M0144. Stage 4 has been split into two distinct steps; rather than the composite view that was shown in version 2.2
  • M0146. Stages 1 and 2 were incorrect and have now been fixed.
  • M0148. The stage reaction mechanism image for stage 7 was incorrect and has now been fixed
  • M0150. The overall reactants and products are incorrect and should be ATP + diphosphate-nucleoside -> ADP + triphosphate-nucleoside
  • M0159. Images for the substrate and products in stages 1 and 2 were incorrect and have now been fixed.
  • M0197. The function of Thr234 has been revisited, and the steric role function removed. It is described as possibly being a metal ligand, with electrostatic stabilising functionality. The H-bond donating interaction with one of the catalytic lysine residues has also been added. Asp197 has been added and we have also included a comment upon the mobility of Lys82 and Arg86 in the active site.
  • Cofactor annotation has now been added to. M0134, M0135, M0137, M0139, M0147, M0156, M0183

Finally, we have identified the issue with the animated reactions in Internet Explorer, and this has now been fixed, although there are still issues with Firefox and Safari in displaying the animations


Version 2.2

Version 2.2.8

This version (17th October 2007) represents the addition of 11 new entries. It also represents an update in the web interface, in which the mechanism and mechanism components also have the mouse-over properties that were introduced for the amino acid residue functions and the entry pages are automatically generated on the fly. Also listed are other MACiE entries that are homologues or have the same EC number to the current entry. The annotated non-metal cofactors and their function are now also included in the entry display. Finally, we have also explicitly linked MACiE entries to the corresponding Metal MACiE entries at the overall reaction level as well as the step level.

Some specific changes have also been made to existing entries:

  • M0001. Added two new references. 11371180 and 10194325
  • M0002. Added the oxyanion stabilising residues (main chain amides of Ser70 and Ala237)
  • M0003. Updated image for step 2 to still include the NAD+ product. Added extensive comments to all steps. Added two references (PMID:11035252 and PMID:11340659) which suggests that the proton may come from the His161, which is in contact with solvent and thus is readily re-protonated and give further support for the enol form of the FAD respectively. Step 4 has been altered to reflect reference 2.
  • M0005. Changed PDB code to 1WHS, to represent an APO state of the enzyme, with a higher resolution than the previous (1BCR) representative code.
  • M0009. Removed potassium ion from the reaction as it is not involved in the stabilisation of the negatively charged intermediates, but in the maintenance of the active site geometry (PMID:14967023)
  • M0010. Representative PDB code for the complete reaction has been changed from 1MKA to 1MKB. 1MKA represents an inactive and covalently modified structure, whereas 1MKB represents the native state of the enzyme.
  • M0020. Added new reference (PMID. 12010043), added cofactor (FAD) and created a new first step in which the FAD cofactor is activated.
  • M0021. Changed EC number from 1.1.1.39 to 1.1.1.38, which is the correct EC number for the PDB code cited.
  • M0023. Species corrected, the PDB structure is of Rat liver glycine N-methyltransferase expressed in E. coli. not E. coli glycine N-methyltransferase.
  • M0058. PDB changed from 1AB8 to 1CJU, which contains the catalytic metal ions required for enzyme activity.
  • M0072. Changed product of step 02 and substrate of step 03 to the ene-diolate, which is suggested to be the correct form of the intermediate due to the fact that it is well stabilised by the Zn(II).
  • M0084. Removed Mg from the annotation as it appears to be simply binding and holding the triphosphate group in place.
  • M0098. Changed EC code from 3.5.1.31 to 3.5.1.88 which is the correct EC code for the reference and PDB code used. Changed name from formylmethionine deformylase to peptide deformylase. Changed PDB code from 1BS4 to 1BSZ, which contains the correct catalytic metal (Fe).
  • M0103. Added reference PMID:10585424, added His422 annotation, as the His422 is essential for activating (by increasing the redox potential of the) flavin.
  • M0105. Added reference PMID:8799115.
  • M0113. Added Cys315 and Lys348 to the reaction, also added new reference PMID:16878980.
  • M0114. Added Cys30 to the reaction.
  • M0115. Added reference PMID:14965342

Version 2.1

Version 2.1.7

This version incorporates Metal Cofactor annotation information from the Metal-MACiE Database (Metal-MACiE: a database of metals incolved in biological catalysis. C. Andreini, I. Bertini, G. Cavallaro, G. L. Holliday and J. M. Thornton. Bioinformatics, 25, 2088-2089, 2009. Medline ID. 19369503). It also represents some minor error corrections from the version 2.0, the changes are as follows:

  • M0004. Changed pdb code from 1nid to 1nia, as the latter contains the biological unit, a homotrimer. The overall reactions now matches the reaction description as per the EC number.
  • M0006. Tyr197 removed as its only role was to move in and out the active site to allow NADPH to bind to the enzyme.
  • M0013. TTQ is no longer considered a cofactor is now treated as two PTM residues. Trp57L and Trp108L.
  • M0023. Removed Asp70A Leu136A Ser139A as these were binding only residues.
  • M0032. Now we depict the reaction mechanism with the R enantiomer of the substrate where two bases are involved (Glu99 and Glu172) as opposed to only one base for the S enantiomer (Glu172).
  • M0035. Added a second Mg2+ cofactor, which is required to activate the enzyme.
  • M0038. Changed pdb code from 1gtp to 1fbx as this structure contains the Zn2+ cofactor.
  • M0041. Glu39 has been removed as it was metal binding only.
  • M0042. Changed pdb code from 1smn to 1ql0 as this structure contains the Mg2+ cofactor.
  • M0053. Removed step 2 as this was a rotation of the substrate in the active site.
  • M0056. Removed step 2 as this was a rotation of the substrate in the active site.
  • M0064. Removed step 2 as this was DNA Uncoiling.
  • M0066. The proton transfer of the second step of the reaction has been moved to the first step, in accordance to PMID. 9538014.
  • M0074. Added a second Mg2+ cofactor, which is required to activate the enzyme. Changed pdb code from 1dae to 1bs1 as this structure contains the Mg2+ cofactors.
  • M0082. In steps 1 to 4 added Arg26 main-chain carbonyl as an activator of the N-terminal amino group, and Trp74 main-chain amide as stabiliser of the nucleophilic cysteine.
  • M0083. Changed pdb code from 1ae7 to 1l8s as this structure contains the Ca2+ cofactor. Changed SwissProt and Species accordingly.
  • M0103. Removed His422 as a catalytic residue as it is a cofactor binding residue only.
  • M0111. In steps 1 to 4 added Arg31 main-chain carbonyl as an activator of the N-terminal amino group, and Phe207 main-chain amide as stabiliser of the nucleophilic cysteine.
  • M0113. Removed Cys315 as a catalytic residue as it is a cofactor binding residue only.
  • M0114. Removed Asp267A as it is mentioned as part of an active site triad, but its function as a catalytic residue is not supported in the literature.
  • M0116. Removed Gln132 (binding only) as catalytic. Added Arg127, Asp135 and Tyr235 as they block access of water to the active site. Proton transfer initiates hydride transfer as opposed to being a consequence of.
  • M0117. Removed Trp677 as a catalytic residue. Unidentified acid/bases assumed to be hydroxonium ions/water in all cases. Final electron acceptor changed to generic hemoprotein. Proton relay pathway in step 3 has been assumed to be activated by water.
  • M0118. Added His256D as part of the proton relay (to relay the proton from the substrate to solvent). Gln228D is charge stabilising the intermediates of the reaction and was also added. Formation of the urate hydroperoxide has been split into two steps via radical intermediates.Changed pdb code from 1r56 to 1wrr which is of higher resolution.
  • M0119. pdb code changed from 1kek to 2c3m. Thr31 and Arg114 have been added as charge stabilisers for pyruvate and CO2. The formation of the radical intermediate is now depicted as one step, as opposed to three steps, due to new evidence PMID. 16472741. The unidentified base has been replaced by a water molecule. The return step has been moved to the end of the reaction. Electron transfers to ferredoxin are now depicted in the same step where the electrons are generated rather than in the last step.
  • M0120. Proton relay from water is now depicted as one step, rather than two. Base-H has been replaced by X00022.
  • M0121. Although Cys185 was not deemed a catalytic residue, it appeared in the diagrams as a metal ligand. In the new version we have removed Cys185 form the diagrams.
  • M0122. Changed pdb code from 1ff3 to 1fva (the latter is of higher resolution and doesn't contain modified residues). Cys218 (Cys198 in 1ff3) is depicted as the proton donor in the dehydration step (Step 3) rather than an unknown base as proposed in PMID. 10841552. Split the reduction by Thioredoxin into two steps. Added an inferred return.
  • M0123. First step (a two electron transfer) split into two single electron transfers. Added Trp234A as catalytic as it acts as a stabiliser of one of the intermediates of the reaction. A new reference by the same group PMID. 16503650 was added and His398A is not longer in its charged state, and therefore it is not acting as a proton donor in the last step, the acid for the return step is now a hydroxonium ion.
  • M0124. Trp172 replaced by the correct residues for the species. Trp126. Added Asp91A,Ser156A and Ser157A as part of the D proton pathway. Added Ser255A,Thr316A and Lys319A as part of the K proton pathway. His240A added as is cross-linked to Tyr244A and affects its pKa. All steps involving protons from the D pathway include proton relay through water molecules and the residues Asp91A,Ser156A and Ser157A. Step 10, which is the only step involving protons from the K pathway include proton relay water molecules and the residues Ser255A,Thr316A and Lys319A. Removed step 12 as this corresponded to the de-coordination of the product water molecule and coordination of molecular oxygen. This is now mentioned in the comment of step 11.
  • M0126. The acceptor of the electron in the first step, and subsequent donor in the fifth step is assumed to be the closest iron-sulfur cluster (FS4267). The unidentified acid giving a proton in step five is assumed to be the hydroxonium ion. The unidentified bases in step six have been replaced by water molecules.
  • M0128. Due to lack of evidence suggesting an enzymatic base we have assumed that the unidentified base in step 2 is a water molecule.
  • M0131. Proton relays now begin with a solvent water molecule rather than His72, so that the relay produces an hydroxonium ion instead of a protonated His72. Added Pro293 and Lys297 as catalytic residues. The former prevents the product, which should be more reactive than the substrate, from exposing its 5-carbon to attack in subsequent cycles of catalysis and inhibits formation of the out conformation for enzyme reduction in the presence of product. Pro293 is also known to be fundamental to the conformational response of the protein to signals from the substrate pOHB. Lys297 stabilises the flavin alkoxide leaving group in step 6 (PMID. 15568817).
  • M0132. Formation of the 4a-hydroperoxy-FMNH is depicted now in 2 steps rather than 3, i.e. the proton transfer of step 3 has been added into step 1. Step 5 has been split into two steps. homolysis of the O-O bond and electron transfer plus a second step for the proton transfer from the intermediate to the His44. Added His45 as a catalytic residue that stabilises the formation of the first two intermediates of the reaction (PMID. 10194361).
  • M0133. Evidence suggest (PMIDs 15238007 and 17017800) that several steps are concomitant with each other. Therefore steps 3 and 4, 5 and 6, and 7 and 8 have been joined together. Evidence (17017800) suggest that the Glu366 proton channel is less plausible than the Asp251 channel, as the former residue is buried in the protein and no proton source is close to it, whereas Asp251 is connected to the protein surface by a single residue. Arg186. Therefore we have removed Glu366 as the proton source and replaced it by a proton relay through Arg186 and Asp251.
  • M0136. Trp62, His146 and Asp228 have been added as Electron Relay residues. Thr199 was added by its implication in oxygen activation. The two electron transfer was previously depicted as a one step reaction, but now it is stepwise.
  • M0137. The formation of the planar peroxo adduct is now shown as two steps rather than just one. Step 3 was just change of substrate, i.e. a non-catalytic step and has been removed. BondInvolved added in Overall Annotation.
  • M0138. This five step reaction has been reduced into two catalytic steps. Steps 1 and 2 have been joined together, as have steps 3, 4 and 5. Changed pdb code from 1cbj (Bos taurus) to 2jcw (Saccharomyces cerevisiae).
  • M0139. Added Glu802 as an electrostatic stabiliser. The unidentified base has been replaced by a water molecule and the unidentified acid by a hydroxonium ion. The nucleophilic group in step 3 has been replaced by an hydroxide instead of a water molecule, and therefore there is no need to deprotonate it.
  • M0140. Hydrogen transfer from/to the Cys408 to/from the cofactor is now depicted as two steps. homolysis/colligation + hydrogen transfer. Split the reduction of disulfide bonds into two steps.
  • M0141. pdb code 1wvf replaced by 1dii. Added Ala649C and Met650C as electron relay residues. Acceptor has been replaced by electron carrier protein (PMID:10623531). Steps 1 and 2 were joined together to form a proton relay through Tyr473A. Likewise, steps 3 and the inferred return step (step 13) were also joined together.
  • M0142. We have added a new reference in which the electron transfer is said to occur to an Adrenodoxin dimer. The first electron is transferred to the non-bound Adrenodoxin molecule which then dissociates. Then a second electron transfer occurs to the remaining Adrenodoxin molecule.
  • M0143. Added Cys10 MainChainAmide, Thr11 MainChainAmide, Cys15 MainChainAmide and removed Ser17 MainChainAmide as part of the oxyanion hole. PMID 16797027 and 16905151.
  • M0144. The additional oxygen ligand is considered now an elongated oxo group rather than a hydroxyl group. This avoids the need of an unidentified base in step 2, however it means an additional step (step 4) with a new unidentified base.
  • M0145. In the first step, the base was replaced by one of the water molecules coordinated to the iron centre. The second cyclisation (old steps 5 and 6) is now depicted as an heterolytic one step process rather than a radical two step reaction. Added an inferred return step to regenerate Fe2+ coordination sphere. Added Phe211 as a catalytic residue. BondInvolved added in Overall Annotation.
  • M0146. Removed Ser175A as this was a metal ligand residue only. Step 8 has been split into two one electron transfers in analogy to other molybdenum containing enzymes within the database. Water molecules are used as the acids/bases needed for the rearrangements in steps 5 to 7.
  • M0160. Removed the two water molecules ligated to the Mg2+ ion. Corrected the first step to match the actual products of the reaction.
  • M0161. Added Glu65 as a Lewis acid that activates the phosphate group. Added an inferred return step.
  • M0162. The glycosylase step is now depicted as an SN2 reaction mechanism, and no as an SN1 (two-step reaction). The unknown base has been assigned as Glu23. Arg22A and Arg26A have been added as stabilisers of the basic leaving group.
  • M0163. Removed Glu48 and Asp134 as catalytic as they are binding the water molecules only. An inferred return step has been added to return His124 to its native state.
  • M0165. Added an inferred return step
  • M0166. First step divided into two as references claim it to be a SN1 reaction. Added an inferred return step. CATH code 1.25.40.70 has been removed.
  • M0168. His152 is depicted now in its protonated form. Gly291 has been removed as catalytic as there is no support from the authors for this residue as being catalytic.
  • M0169. 1tk3 has been replaced by 1pfq as the later one is of higher resolution. First step has been split into two. an activation step and then the nucleophilic attack of the Ser, as for other Ser-His-Asp triads in MACiE. Asp now remains negatively charged throughout the reaction, with His becoming protonated.
  • M0170. Added His513 as catalytic residue. First split step into two.
  • M0171. First two steps are now just one. Added an inferred return step.
  • M0172. The previous pdb code (1ybq) correspond to a mutant enzyme, so we have changed the pdb code to 1onw.
  • M0173. Third step divided into two, as reference states the reaction occurs in an SN2 fashion, but through a tetrahedral intermediate state.
  • M0175. Thr26 MainChainAmide was replaced by Gly27 MainChainAmide in the basis of quantum chemical studies supporting this residue as the stabilising ones PMID:8551523.
  • M0176. Added Tyr157 as a HBD and steric regulator of the water molecule and Glu143 as an electrostatic stabiliser of the Zn cofactor. Added a third step in which a proton is transferred from the C-terminal residue of one of the product proteins to the N-terminal residue of the other product protein.
  • M0177. Added Arg19 main-chain carbonyl as an activator of the nucleophilic threonine. Asp17 is now depicted negatively charged as in ref 2 (PMID. 9087403).
  • M0178. Added Arg373C as a Electrostatic/Charge Stabiliser.
  • M0179. Changed pdb code from 1a6d to 1q3s, therefore residues Asp60, Thr97 and Asp393 are now Asp64, Thr98 and Asp393. Added side chains of Thr98 and Thr97 as these stabilise the negative charges of the gamma phosphate. Also we now depict Asp393 as a proton acceptor/proton donor.
  • M0180. Changed pdb code from 1qco to 1hyo. First step split into two as we consider the transition state intermediate a real intermediate of the reaction. Added an inferred return step. Glu364 depicted as charge stabilising the His133 rather than abstracting a proton from it.
  • M0182. Changed pdb code from 1oqf to 1mum. Added inferred return step.
  • M0183. Added two steps to photo activate the FADH-. Added three catalytic residues. Trp277, Glu275 and Asn341. The first involved in electron transfer between FADH- and CPD. The second two are involved in intermediate stabilisation.
  • M0184. Added an inferred return step
  • M0185. This reaction was replaced, as it was performed by the same enzyme that performs M0162. The new pdb code corresponds to 1lwy.
  • M0186. Added Cys303 as this residue is an electrostatic stabiliser of the PLP cofactor. In steps 4 and 5 the unknown base has been replaced by the phosphate group of the PLP in analogy to its role in steps 1 and 3. Changed pdb code from 1pwe to 1pwh as the latter contains the cofactor.
  • M0187. Changed pdb code from 2mnr to 1mns as the latter contains the real cofactor Mg (as opposed to Mn in 2mnr).
  • M0188. Added Lys153 as a stabiliser of the negative tyrosine.
  • M0189. Added His126 as it provides additional stabilising interactions with the protein substrate.
  • M0190. Changed pdb code from 1hzt to 1nfs as the latter contains the metal cofactors. Added Tyr104A as the proton donor of the reaction (before it was Glu116A) going through a proton relay by Glu116A. Added an inferred return step.
  • M0192. The water molecule is now depicted as an hydroxyl ion due to its stabilisation by Tyr107. Added an inferred return step.
  • M0193. Ser17 has been removed as a catalytic residue due to lack of evidence for its role in the literature.
  • M0195. Added Asp245 as an electrostatic/charge stabiliser of the PMP cofactor. The 3 Schiff base formations and the last water addition have been now reduced to 2 rather than 3 steps, making a reaction of 12 catalytic steps rather than 16. By analogy to aspartate aminotransferase, the unidentified base has been assigned as Lys273.
  • M0196. Removed Lys97 and Lys109 as catalytic residues, as theoretical evidence for the residues contradicted experimental results also cited for the entry.
  • M0197. Added Thr40A, His45A, His48A, Lys82A, Arg86A, Lys230A, Lys233A and Thr234A as catalytic residues. They stabilise the transition states of the catalytic steps of this reaction.
  • M0198. Removed the unidentified base on the last step and changed the ionisation state of the CoA substrate from neutral to negatively charged.
  • M0199. Removed Lys125, Gly166 and Gly167 as they are binding residues only. Removed Thr288 as its role of charge stabiliser of the second intermediate of the reaction is not supported in the literature. Added Lys160 as a charge stabiliser of the intermediates of the reaction. Replaced the unidentified base in the last step and now use the phosphate leaving group as the proton acceptor.

Version 2.0

Version 2.0.6

This expands coverage in MACiE to 202 entries, and is officially released in the 2007 Database Issue of NAR (MACiE (Mechanism, Annotation and Classification in Enzymes). novel tools for searching catalytic mechanisms. G. L. Holliday, D. E. Almonacid, G. J. Bartlett, N. M. O'Boyle, J. W. Torrance, P. Murray-Rust, J. B. O. Mitchell and J. M. Thornton. Nucleic Acids Research, 35, D515-D520, 2007. Medline ID. 17082206).


Version 1.1

Version 1.1.6

This version represents an updating and fix of the first version of MACiE. It also includes the following website updates:

  • "Curly arrows" added to reaction steps
  • Added UniProt code search to MACiE
  • Added a catalytic domain search option to the CATH code
  • Added an overall comment query
  • Amino Acid Query updated to allow up to 6 residues to be queried for with up to 2 function each. This query replaces the individual and combination queries previously available.

Where appropriate inferred steps for returning the enzyme into its native catalytic state have been added. The only reactions which do not return and do not have a return step in MACiE are:

  • M0008. this mechanism uses unidentified bases, and so cannot be said to return
  • M0037. the return step is more than a simple proton transfer, and the references in used are unclear as to the mechanism of the return
  • M0038. the mechanism of the last step is unclear, and thus the return step cannot be clearly postulated.
  • M0070. this mechanism uses unidentified bases, and so cannot be said to return

Reactions updated from Version 1.0:

  • M0001. The reactant and products were incorrectly named, this has been fixed.
  • M0003. Unidentified base removed in favour of a "free" proton which has come in from outside the enzyme active site. The reference is clear about this, but not how the proton gets in in the first place.
  • M0004. Residue His255 is now His255' as the catalytic residues are from different chains.
  • M0006. Updated with a new PDB code. 3GRS. The original PDB code was 1GET, which was from an engineered protein, which accepted NAD rather than NADP. The mechanism has also been corrected, which is from a new reference that has also been added.
  • M0007. Missing reference added, Biochem 1991, 30, 8671-86788, PMID=1888729. Final step annotated as an inferred return as a direct proton transfer between the Asp283 and Lys230.
  • M0009. Changed to reflect newer reference (by same authors as our original reference) with an updated mechanism and to change the amino acid residues listed as we previously had residues which were ONLY binding and metal ligands, not doing any catalytic activity. Changed PDB code to 1P7L, which is a higher resolution structure with substrates bound. Return added to step 1 because this is the only step in which the enzyme is actively involved. MACiE also uses an updated reference. PDB was 1FUG.
  • M0010. Added Val76 main chain carbonyl to MACiE which is acting to make the neutral His70 more basic.
  • M0011. We have changed this reaction so that it is a single step. rather than two steps, as the product of the first step in version 1.0.6 was a transition state.
  • M0013. Added extra step (1) as methylamine's pKa known to be 10.6, and so requires a base to deprotonate it before it will react. This is tentatively identified as Asp76. Steps 4, 7, 11 and 12 have bases removed (i.e. are assumed to be non-base catalysed, although this may not be the case). The base in step 6 is assumed to be Asp76 from alignments with Quinohemoprotein Amine Dehydrogenase (ref Davidson 2005) Asp32 is also assumed to be a base in the reaction step 8. Inferred enzyme regeneration steps have been added (steps 5 and 13). Step 5 is assumed to be the product water molecule deprotonating Asp76, and step 13 is assumed to be the ammonia (-> ammonium) and a water deprotonating the Asp76 and Asp32. Trp108 and Trp57 (aka Trq57) have been removed from the annotation, and are now only listed in the comment of the overall reaction. This is because they are post-translationally modified and part of the cofactor.
  • M0015. Added Asn193 to bring into line with CSA entry, removed Asp103 which only seems to be a metal ligand.
  • M0016. Added regeneration step in which the "product" is protonated and the OH- is regenerated.
  • M0017. Substrate 1 was incorrectly drawn. This has been fixed. Glu201 seems to be binding only and so has been removed from the mechanism. Substrate in reaction steps 2 and 3 was incorrectly drawn, this has been fixed.
  • M0018. Added an inferred return step with water acting as the base. Water has been chosen as there is no evidence for another amino acid residue being involved and the enzyme is in an aqueous environment. We have also added the residues Asp305 and Asp170 which are interacting with the substrates to make the carbon of the C=N bond more reactive. Changed pdb id from 9jdw to 1jdw.
  • M0019. Added Ca2+ cofactor. Added inferred return step using a proton relay through waters in the active site between the Asp275 and Glu132 (which are approx 10 angstroms apart). Added Arg136 which affects the pKa of the Glu132.
  • M0020. Third step of the reaction is removed as the reference states that the re-protonation of the substrate is via a proton relay through Arg381, Glu378 and Arg402, whereas previously we had a two step mechanism for this proton transfer.
  • M0021. Asp278 added to activate the Lysine to act as a base. Inferred return step added, with proton relay occurring through the Lys183. The distance between Tyr112 and Asp278 is 5.6 Angstroms, the distance between Asp278 and Lys183 is 3.4 Angstroms and 2.9 Angstroms between Lys and Tyr.
  • M0022. Added an inferred return step with water carrying out the return. Water is assumed to be carrying out the de-protonation.
  • M0023. Removed Ala13' and Trp30' from the mechanism as they appear to be binding only. Added Asp70, Leu136 and Ser139 as there appear to be important to the stabilisation of charge formed during the reaction.
  • M0024. Added Trp137, which is involved in intermediate charge stabilisation, and Ala86 which is involved in the activation of the His90 (in a triad-like set-up).
  • M0026. Changed main mechanism from 3 steps to 1, and added an inferred return step. Added 2001 Biochemistry reference. Added Asp33.
  • M0027. Removed Glu4, Phe66, Tyr56 and Asn134 as these seem to be binding only. Added an inferred return step.
  • M0028. Removed Glu341. Reaction changed from 5 steps (which included transition states) to 2 steps
  • M0030. Added Trp333, which stabilises the Cys418 radical, and Reference to 1999 Structure paper.
  • M0031. Added an inferred activation step of the tetrahydrofolate reactant, this is necessary to get the reactant into the correct conformation. No direct support for the mechanism shown was found, but it is based off evidence in the references used. Added 2 new references (medline 97263753 and 98301389). Added comment in steps 1 and 2 to specify the role of Arg218 which, from the diagram, is not very clear.
  • M0032. Added a spontaneous first step, which probably does not occur inside the enzyme. This step is necessary to generate the substrate that the enzyme actually works on. The overall reaction has been updated to reflect the EC code, however, the enzyme only works on the product of the spontaneous reaction between the overall reactants. Corrected entry to reflect residues belong to different chains.
  • M0034. Unidentified base removed from step 5. Added residues His246 and Tyr255 to reflect entry in the CSA. Added an inferred regeneration step to protonate the product and regenerate starting state of the iron cofactor.
  • M0035. EC changed from 2.7.1.38 to 2.7.11.19
  • M0036. Added an inferred return step.
  • M0037. A return step cannot be added for this reaction as there is no evidence in the references used to support a mechanism by which this happens, and the return require more than a simple proton transfer.
  • M0038. The actual mechanism of the final step of this reaction is unknown, and thus no return step can be added, as the function of the amino acids in the final step is also unknown.
  • M0039. Added an inferred return step.
  • M0041. Added an inferred return step of a proton relay through water as the distance between the two Histidines involved is approx 5.7 angstroms.
  • M0043. 1VHR has been replaced by 4KBP. The entry for this reaction in the original database was an incorrect mechanism for the PDB code, and we had originally changed the PDB code rather than the mechanism. This was deemed the wrong thing to have done and so we have now altered the entry to reflect the correct PDB code and mechanism.
  • M0045. Arg227, His327, Asp328 which are involved in transition state stabilisation, have been added to the MACiE entry. Changed PDB code from 1CGT to 1CDG.
  • M0046. Base identified as water.
  • M0047. Inferred return step added as a direct proton transfer between the Glu290 and Asp356. Arg409, which is involved in Transition state stabilisation, has been added to the MACiE entry
  • M0048. Added an inferred return step in which the proton is given up to water. Added residues Asp134 and Glu133, which are acting as transition state stabilisers, to bring into line with the CSA. Added reference. PMID 9521733
  • M0049. Adding Tyr62, Ser81'', Glu66, and Glu197'. Renaming Phe83' to Pvl82'. Bases identified as Glu197' Ser81'' and Tyr62. Added an inferred return step. Added references 2197977 (Annual Rev. Biochem.) and 8490030 (Biochem)
  • M0050. Inferred return step added, with the proton coming from an active site water molecule. Comment added to the first step. the references for this reaction state that the mechanism is an electrophilic substitution, however, the substituting group is a proton. By our definition, a proton cannot be a substituting group, and thus in MACiE this reaction is termed an unimolecular elimination from the conjugate base and a proton transfer. Changed chain of residue Asp65.
  • M0052. Arg331 has been removed as it is just a phosphate binding residue. An inferred return step has been added, this is a proton relay through a water molecule as the shortest distance between the Asp and Glu is 6.95 Angstroms. Added reference PMID 9878448, which cites the rate determining step as step 2 (the carbon-carbon bond forming step).
  • M0054. Concatenated steps 8 and 9, which makes more sense in terms of proximity of the reacting atoms. Also amended mechanism in accordance with a new reference (2) added, which states that although the His is acting as a general acid/base, it is working through the Glu due to the low pKa of the His.
  • M0055. Mechanism in MACiE version 1.0.1 is incorrect. New evidence found for a "correct" mechanism. An inferred return step has been added in which water deprotonates the Tyr and a water is responsible for the return of His and Glu to their native states. PDB code changed to 1GU1. Residues now. Pro15 Asn16 Arg23 Try28 Asn79 Ala82 Glu104 His106 Arg113
  • M0056. Version 1.0.1 has an incorrect EC code. Should be 5.4.99.12. PDB code updated to 1ZE1 from 1DJ0. Added references. 11976723 (Nat Struct Biol 2002) and 16300 (Biochemistry 2005). The reactants and products have been corrected (should be tRNA Uridine and tRNA pseudouridine, not "plain" uridine.) Mechanism corrected to reflect the newer references, a tyrosine has been added as the general acid/base in the final step.
  • M0057. Removed Cys109 as metal binding only. Removed Cys112, Ser113 and Cys114 as these are only involved in binding the NO in the inactive form of the enzyme. Added Arg56 as the unidentified base
  • M0058. Removed Asp 440 as this appears to be a metal ligand only. Altered mechanism so that the substrate is performing the proton abstraction in the first step. PDB code changed from 1AZS to 1AB8.
  • M0059. Added Arg259 as a general acid/base in the lats two steps of the reaction. Other general acid/bases are identified as water. Also added are Lys250, Arg294 and Asn268 which are important in preventing epimerisation of the substrate in steps 4 and 5.
  • M0060. His 143 now annotated as being involved in the decyclisation (step 1), and cyclisation (step 6) through a proton relay. Step 2 is split into 2 steps, reference states that the cis-enolate-ammonium is an intermediate and Asp72 is the general acid/base involved in this tautomerisation. Added reference Biochemistry. 2001 Aug 28;40(34):10187-96.(11513596 ) which states that the reverse reaction binds the linear form of the product, therefore the product of the enzyme reaction is the linear, not cyclic form and thus the cyclisation occurs spontaneously outside the enzyme. PDB changed from 1cd5 (which is the inactive T state of this allosteric enzyme) to 1dea which is the active R state of the enzyme with 2 phosphate ions bound.
  • M0061. Arg11 removed from MACiE as it appears to be simply binding. This brings MACiE into line with the CSA
  • M0062. Added Asp608 and Lys604 which form a triad with the His610 which stabilises the cobalt in its free form. This brings MACiE into line with the CSA
  • M0063. reactants and products were the wrong way around, fixed. Added reference PMID 10467146 which states the His16 is binding and stabilising the Co(II).
  • M0064. Unidentified base removed, reference 12007989 added to support the stabilisation of the tyrosine in it's phenolate form. MACiE PDB id updated from 1i7d (which has the catalytic Tyr mutated to Phe) to 1d6m (which is wild type and also included in the CSA). Reactants and products renamed to single-stranded DNA (rather than generic DNA).
  • M0065. Added a water as a proton relay medium into the inferred return step.
  • M0066. Added in missing references PMID=9749913 (Protein Eng. 1998 Aug;11(8):613-9) and PMID=9538014 (Biochemistry. 1998 Apr 7;37(14):4958-67). Changed transaldimination steps from 3 to 2 (as suggested in the reference). Added in Leu201 as a catalytic residue, which keeps the PLP in the correct conformation, also added is Tyr31, which stabilises the Lysine 145.
  • M0067. Arg15 removed as simply binding the substrate carbonyl. Tyr93 has been added as a steric inhibitor which affects the stereochemistry of the reaction. However, the function of this residue is only tentatively assigned. Added inferred return step in which ammonia acts as the base to deprotonate the Glu117.
  • M0068. Added reference PMID=7601336. Added in residues main chain amide of the Glu254, also added are Ser136 and Met135, both as main chain amides, which are stabilising the negative oxygen of the FADH-. It should be noted that these two residues have been inferred from the PDB structure, and not from a direct citation in the references used. Added in two steps in which the FADH- is reduced back to FAD by two single electron transfers to electron-transferring flavoprotein (ETF). An inferred return step has also been added. Overall reaction has been updated such that the FAD is now a cofactor only, and ETF (x2) is listed in the reactants and products, and 2 protons are also produced.
  • M0069. Added reference PMID=15044493, which states that in the original PDB deposition, the His was in a different (and incorrect) tautomeric form and is now stabilised by Asp126. We have thus added Asp126 into MACiE.
  • M0071. Added residue Gln63 added in as an electrostatic stabiliser, residue identity inferred from the human form of this protein (1emh). Also added were side chain amide of Asn123 and main chain carbonyl of Asp64, which are stabilising the flipped out conformer of the uracil. Added inferred return step. Added a step (which occurs outside the enzyme spontaneously) to generate the uracil from the tautomer formed from the reaction.
  • M0073. The references in MACiE stated two possibilities for the mechanism, further investigation has offered greater support for one over the other, and the mechanism in MACiE has been updated to the preferred mechanism. We have added His63, which is acting as a general acid/base in the mechanism, also added are the main chain amides of Glu81 and His80, which are stabilising the negative charge on the substrate, and the side chain of Glu58 which is in a dyad with the His63.
  • M0074. Glu12 is being removed from MACiE as the reference states it is binding only.
  • M0075. Added inferred return to the final step of the reaction. The leaving AMP is reprotonated from the Asp46.
  • M0076. Added inferred return to the final step, in which the leaving nicotinamide ring is reprotonated from the Glu988.
  • M0078. Hydrolysis step is now assumed to be a single step, proceeding through a tetrahedral transition state. Added reference PMID=8718855 and J Phys Chem B reference (DOI. 10.1021/jp034717). Added Ser244 which is stabilising the neutral His274.
  • M0079. Added return as part of the final step, as Glu317 acts in a proton-relay type manner.
  • M0080. Added an inferred return step which is two proton transfers involving two water molecules as the distance between the catalytic histidines is 8.18 Angstroms. Substrates in reaction step has been altered to mirror the substrate in the overall reaction.
  • M0081. Added reference PMID=7479847.
  • M0082. Changed PDB code from 1moq to 1jxa (which is the complete enzyme with both domains present. 1moq is the isomerising domain only). Changed the inferred transaldimination and Schiff base formation steps from 3 to 2 steps. Added Lys485 in a general acid/base capacity and Glu481 which is "activating" the Lys485 through H-bonding interactions. Added reference PMID=11700065 for structure 1JXA.
  • M0083. Main chain carbonyl of Gly30 removed from MACiE as it is a metal ligand only.
  • M0085. Removed Lys23, Ser65, Thr45 and Arg150' as these seem to be binding the phosphate only. Added Asp91 (stabilises the positive charge on His19), Asp101 (destabilises Asp71) and Arg107C (stabiliser negative charge on Asp101).
  • M0086. Removed Glu314 which is an alternative base. Either Glu223 or Glu314 may act as the base in this active site, the mechanism is identical, whichever one is acting, and we only list one of the mechanistic possibilities. Added an inferred return step, in which the liberated phosphate (of ADP) takes the proton off the Glu225 through a proton relay through a water molecule, which is coordinated to the Magnesium ion, and is H-bonded to both species.
  • M0087. Added Asp221 and Arg336 which are affecting the pKa of the His320. Added reference PMID=8718850.
  • M0088. Added His164 (main chain carbonyl and side chain) which is positioning and stabilising the His166, His166 main chain carbonyl has also been added which is holding the catalytically important water (see Asn153). Also added are a Zn2+ cofactor and Asn153. which is stabilising phosphate through a water molecule.
  • M0089. Added in residues Phe76, Phe77, Asn219 and Trp308 which are all responsible for stabilising the carbocations formed during the course of the reaction. Phe77 and Asn219 are also responsible for forcing the substrate into the correct conformation. Added an inferred return step where the His309 is deprotonated by a water molecule.
  • M0090. Added to MACiE His54 and His300, which are involved in the addition of water. Added an inferred step which regenerates the His54 and produces the protonated C00155 (L-homocysteine). Also added are Asn190 and Ser360. Added reference PMID=11927587. Removed Glu155 which seems to be binding only.
  • M0091. Added an initial step in which the His82 deprotonates the phosphate before the uridine binds.
  • M0092. Changed mechanism such that the general acid/base is the structurally conserved water molecule, activated by Asp264. Changed final E-S bond cleavage proceeds via a tetrahedral intermediate (which is stated in the reference) and an inferred return step has been added. Lys204 assumed to be protonated and Cys260 is negative (pH is 9).
  • M0093. Added an inferred return step in which the His866 and Glu559 both deprotonate a hydroxonium molecule
  • M0094. Removed proton relay through His in the Ser-His-Asp triad.
  • M0095. Added reference PMID=2304904. Added an inferred return step.
  • M0096. Added an inferred return step and a step which regenerates the His for attack of the hydroxyl group. Added reference PMID=1696320, changed current reference PMID to 10387007.
  • M0097. Removed residues Asn89, Glu91 and Thr127 which appear to be binding only, this brings MACiE into line with the CSA. Mechanism corrected.
  • M0098. Removed Ile44, which appears to be binding only. Step 1 is split into two steps, with the proton transfer occurring prior to the binding of the substrate.
  • M0100. Added inferred return step.

Version 1.0

Version 1.0.5

This version change incorporates further user comments on the previous version and extends some of the search functionality.

  • A Homology search has been added to MACiE on the PDB query. This is based off the homology listings in the Catalytic Site Atlas.
  • The EC query will now "walk" up the EC code until it finds a match in MACiE, thus EC 1.1.1.1 (which is not in the currently in MACiE), will try 1.1.1.- as a query, if this was not present, then EC 1.1.-.- will be queried for, and finally, EC 1.-.-.- will be queried for.
  • The font face has been changed from Times New Roman to Arial, to give the web site a more updated look
  • The list by pages have been updated such that all accession codes take the user to the relevant resource, e.g the PDB code links into the RCSB site, the EC number links into the IUBMB site, the CATH code links into the CATH site. The only way to get into the MACiE entry is to click on the MACiE code, which is now always listed first.
  • The Chemistry Query has been updated to include three bond changes in the query. The bond formation/cleavage and order change queries have also been split, due to the difference in how these data are held in MACiE.
  • Added a MACiE FAQ
  • Added a cautionary note to all animations
  • Added a reaction step comment query
  • Overall reactions are now shown in the same order as they are listed
  • Added a listing of all PDB codes which are in the CSA and determined to be homologous to the PDB code in MACiE

Version 1.0.4

This version change incorporates user comments on the previous version.

  • The home page has been updated to include:
    • a direct link to the searches. The link in the top panel has been retained for navigation when deeper in the bowels of the database.
    • a search by enzyme name or partial enzyme name
  • Links from the query results page now open in a new browser window
  • A list of funding bodies has been added to the MACiE home page

Version 1.0.3

This version takes MACiE out of frames

Version 1.0.2

This version represents the development of a searchable version of MACiE online.

  • Added a function to browse for specific MACiE entries by their EC number, CATH code or PDB code. This replaces the original home page and links to the information on how to view (and what is contained in an entry) is now linked from the home page.
  • Added search link to the banner. This links to a search menu. Added some simple amino acid residue queries for individual residues (by specific location of function and activity or by a specific activity) and a combination of 2 residues. This is still under development. A known bug is that if two of the same residue type/activity/location are imputed (e.g. two Asp side chain reactants are searched for), the result returned is incorrect.
  • Added in more generic searches for amino acids, it is now possible to search for an amino acid residue irrespective of location of function, or activity, or both.
  • Added in more generic searches for combinations of 2 amino acids. Function and Activity default set to "either". Added search function for a single amino acid by it's location, activity and function. For example, it is now possible to search for all Tyr residues acting as a reactant, through the side chain as a nucleophile. Function default set to "unknown function".
  • Split combination of residue query and individual residue query. Amino acid residue is now a pull down option. Added combination of function search to individual residue query. Made default options more obvious. all default searches should produce a positive result. Added mechanism query. Currently only contains a search for an individual mechanism, 2 mechanisms and 3 mechanisms occurring in a reaction step.
  • Added query by mechanismComponent to mechanism query. This does not include bond formation, bond cleavage and bond order change, these will be in the chemistry query. A combination of 2 mechanismComponents query has also been added.
  • Added queries for combinations of mechanism and mechanism components. Added queries on bond changes in MACiE. This is an incomplete implementation as the annotation of bonds in MACiE is not always consistent, i.e. C-O may also be annotated as O-C. This is not currently supported by the search. Also added queries by (individual or combinations of) reactive centre. Added queries for chemical compounds (by KEGG compound id) involved in the overall reactions.
  • Added some simple statistics on MACiE. Growth of the database, and frequency of amino acid residues performing a specific function. All statistics are linked from the bottom panel of the home page.
  • Added mechanism statistics. Changed the banner and home page to make the banner smaller. Mechanism and mechanism component pull down lists are now alphabetical. Added query for mechanism and bond type combination.
  • Added amino acid residue and mechanism combination query. Added amino acid and bond type change combination, this is only available for one amino acid at present. Added amino acid residue, bond type and mechanism combination query.
  • Added to mechanism and bond change queries for mechanism component/mechanism combinations. Added mechanism component/mechanism combinations to the mechanism and amino acid query. Added mechanism component/mechanism combinations to the mechanism, amino acid and bond query. Added query by enzyme name (partial matching) and species names (scientific and common names)
  • Added option on home page to retrieve a specific MACiE entry.
  • Added javasciprt to force links to the MACiE frameset. Unfortunately this breaks the back button.
  • Added compound name search to overall reactants and products searches. This is implemented as a partial match to cover as many hits as possible. The names used in MACiE tend to be those found in KEGG. It should be noted that this search is for the overall reaction only.
  • Added cofactor search into MACiE, which searches the substanceList's. Cofactors found in MACiE reaction steps are only likely to include those metal ions which are not involved in the reaction. This is because if a metal ion, or other cofactor, is involved in an overall reaction as a cofactor, and is an active species (i.e. is chemically involved, e.g. PLP) then it will only be included as a reactant. Metal ions which fall into this category will be listed in the reactive centre lists. Metal ion cofactors listed in MACiE are. Mg, Mn, Zn, Ca, Co, Fe, Na, Ni and a generic metal ion (M). Metals which are active species are. Fe. Co and V.
  • Added some more statistics. Graphs of frequencies of amino acid residue functions, by location of function and activity. Graph of reactant vs spectator amino acid residue functions in MACiE. Added in overall statistics on the growth of MACiE by EC number (full and upto the sub-subclass) and CATH code. Added in mechanism component distribution and mechanism component by amino acid graph and data table. Added some bond type statistics, counts of number of bonds (of each type) formed, cleaved and changed in order by reactions and bonds. Also added a total count of bond types formed, cleaved and changed in order in MACiE.
  • Added bond type analyais by amino acids. Added frequency of all amino acid residues, irrespective of function. Added in main chain vs side chain frequencies of amino acid (by activity) and reactant vs spectator frequencies of amino acid (by location of function).
  • The link section has been updated to include IntEnz and ChEBI. Each entry now links to IntEnz
  • Added ChEBI identifiers to the CML version of MACiE, also added in links overall reactant and product molecules in KEGG and ChEBI. Also added in EzCatDB identifiers where appropriate.
  • Added tooltips to the amino acid residue functions, and the home page searching options.
  • Added in ChEBI identifier search on overall products and reactants
  • Added in catalytic propensity statistics based on the method by Bartlett et al. as published in "Analysis of catalytic residues in enzyme active sites", G J Bartlett, C T Porter, N Borkakoti & J M Thornton, Journal of Molecular Biology (2002) Vol 324 p 105-121.

Version 1.0.1

Released December 2005 to coincide with Bioinformatics, 2005, 21, 4315 and based on Version 1.00 of our MACiE dataset (containing 100 entries). The web-version of MACiE 1.0 consists of static html pages and is un-searchable. Entries are browsable through HTML look-up tables, which are arranged by either EC number, CATH code, PDB code or MACiE entry number. Approximately half of the entries have SVG animations associated with them

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