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PDBsum entry 1u5h

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Lyase PDB id
1u5h
Jmol
Contents
Protein chain
223 a.a. *
Ligands
FMT
Waters ×156
* Residue conservation analysis
PDB id:
1u5h
Name: Lyase
Title: Structure of citrate lyase beta subunit from mycobacterium tuberculosis
Structure: Cite. Chain: a. Synonym: citrate lyase beta subunit. Engineered: yes. Mutation: yes
Source: Mycobacterium tuberculosis. Organism_taxid: 1773. Gene: rv2498c. Expressed in: escherichia coli k12. Expression_system_taxid: 83333.
Biol. unit: Trimer (from PDB file)
Resolution:
1.65Å     R-factor:   0.268     R-free:   0.272
Authors: C.W.Goulding,T.Lekin,C.Y.Kim,B.Segelke,T.C.Terwilliger, D.Eisenberg,Tb Structural Genomics Consortium (Tbsgc)
Key ref:
C.W.Goulding et al. (2007). The structure and computational analysis of Mycobacterium tuberculosis protein CitE suggest a novel enzymatic function. J Mol Biol, 365, 275-283. PubMed id: 17064730 DOI: 10.1016/j.jmb.2006.09.086
Date:
27-Jul-04     Release date:   12-Oct-04    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam  
P9WPE1  (CITEL_MYCTU) -  Citrate lyase subunit beta-like protein
Seq:
Struc:
273 a.a.
223 a.a.*
Key:    Secondary structure  CATH domain
* PDB and UniProt seqs differ at 5 residue positions (black crosses)

 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     oxaloacetate metabolic process   1 term 
  Biochemical function     catalytic activity     4 terms  

 

 
DOI no: 10.1016/j.jmb.2006.09.086 J Mol Biol 365:275-283 (2007)
PubMed id: 17064730  
 
 
The structure and computational analysis of Mycobacterium tuberculosis protein CitE suggest a novel enzymatic function.
C.W.Goulding, P.M.Bowers, B.Segelke, T.Lekin, C.Y.Kim, T.C.Terwilliger, D.Eisenberg.
 
  ABSTRACT  
 
Fatty acid biosynthesis is essential for the survival of Mycobacterium tuberculosis and acetyl-coenzyme A (acetyl-CoA) is an essential precursor in this pathway. We have determined the 3-D crystal structure of M. tuberculosis citrate lyase beta-subunit (CitE), which as annotated should cleave protein bound citryl-CoA to oxaloacetate and a protein-bound CoA derivative. The CitE structure has the (beta/alpha)(8) TIM barrel fold with an additional alpha-helix, and is trimeric. We have determined the ternary complex bound with oxaloacetate and magnesium, revealing some of the conserved residues involved in catalysis. While the bacterial citrate lyase is a complex with three subunits, the M. tuberculosis genome does not contain the alpha and gamma subunits of this complex, implying that M. tuberculosis CitE acts differently from other bacterial CitE proteins. The analysis of gene clusters containing the CitE protein from 168 fully sequenced organisms has led us to identify a grouping of functionally related genes preserved in M. tuberculosis, Rattus norvegicus, Homo sapiens, and Mus musculus. We propose a novel enzymatic function for M. tuberculosis CitE in fatty acid biosynthesis that is analogous to bacterial citrate lyase but producing acetyl-CoA rather than a protein-bound CoA derivative.
 
  Selected figure(s)  
 
Figure 1.
Figure 1. Reaction mechanism of bacterial ATP-independent citrate lyase. Citrate lyase contains the prosthetic group, a coenzyme A (CoA) derivative, attached via phosphodiester linkage to a serine residue of the γ-subunit (CitD), which serves as an acyl carrier protein (ACP). CitF, the α-subunit, is the citrate:acetyl ACP transferase which binds citrate and releases acetate as a by-product. CitE, the β-subunit, is the citryl-ACP lyase or citryl-ACP oxaloacetate lyase^3 that carries out the reaction converting citryl-ACP to acetyl-ACP and oxaloacetate. Figure 1. Reaction mechanism of bacterial ATP-independent citrate lyase. Citrate lyase contains the prosthetic group, a coenzyme A (CoA) derivative, attached via phosphodiester linkage to a serine residue of the γ-subunit (CitD), which serves as an acyl carrier protein (ACP). CitF, the α-subunit, is the citrate:acetyl ACP transferase which binds citrate and releases acetate as a by-product. CitE, the β-subunit, is the citryl-ACP lyase or citryl-ACP oxaloacetate lyase[3]^3 that carries out the reaction converting citryl-ACP to acetyl-ACP and oxaloacetate.
Figure 2.
Figure 2. Sequence comparisons of citrate lyase β-subunit (CitE). The secondary structure of Mtb CitE highlights the multiple sequence alignment; red arrows depict the β-strands, the cream cylinders depict the α-helices, the green cylinder depicts α-helix 6a and the yellow cylinder depicts α-helix 6b; the α-helices and β-strand are numbered. Numbers above the sequence correspond to the residue numbers of Mtb CitE. Universally conserved residues are highlighted in blue; residues conserved only among genomes that lack the bacteria α and γ-subunits of citryl lyase are highlighted in pink; and residues involved in forming the trimer interface are highlighted in cyan. Abbreviation of the species names are as follows: M_tub; Mycobacterium tuberculosis, M_mus; Mus musculus, H_sap; Homo sapiens, D_rad; Deinococcus radiodurans, P_aer; Pseudomonas aeruginosa, B_par, Bordetella parapertussis, H_inf; Haemophilus influenzae, S_fle; Shigella flexneri, V_chl; Vibrio cholerae, S_pyo; Streptococcus pyogenes, K_pne ; Klebsiella pneumoniae, E_col; Escherichia coli, M_ext, Methylobacterium extorquens; and the last sequence in the alignment is the enzyme MclA, L-malyl-coenzymeA lyase from M_ext. It is noteworthy that the final six CitE sequences have known α and γ citrate lyase subunits of the bacterial citrate lyase complex. The sequence alignment was made with CLUSTALW and BoxShade. (b) Ribbon diagram of the trimer of Mtb CitE. The structure of CitE was determined by the multiwavelength anomalous diffraction (MAD) phasing method. Three data sets were collected for the SeMet protein in the R32 crystal form at wavelengths near the Se absorption edge at the synchrotron light source at Brookhaven National Laboratories on a charge-coupled device detector. Data were processed using DENZO and SCALEPACK,^18 and MAD phasing proceeded by the standard methods of heavy-atom location (SHELXD [http://www.shelx.uni-ac.gwdg.de/SHELX/]), maximum likelihood phase refinement (ML-PHARE)^26 and density modification [DM].^19 Phase extension to 1.65 Å permitted automated model building for 70% of the protein with ARP/wARP.^20 The remaining residues were traced with the program O.^21 The model was then refined with the program CNS.^22 The geometry of the structure was checked with ERRAT.^23 The α-helices and β-strands are colored as in (a). The Figure was generated using PYMOL [http://www.pymol.sourceforge.net/]. (c) Illustration of the catalytic site of Mtb CitE. A cartoon of the catalytic site complexed with a magnesium ion and oxaloacetate. The ribbon diagram is colored the same as in (b), except the β-strands are colored wheat. The residues that form hydrogen bonds with oxaloacetate and the magnesium ion are shown in a stick representation; carbon, oxygen and nitrogen atoms are colored white, red and blue, respectively. The oxaloacetate is shown in stick form, its carbon and oxygen atoms are colored orange and red, the magnesium ion is shown as a yellow sphere and the water molecules are colored red. The hydrogen bonds are shown with broken black lines and the distance between the atoms is indicated. (d) Illustration of potential binding cavities for citryl-CoA. The illustration shows a ribbon diagram colored as in (c). The blue patches on the surface of the monomer are the conserved charged residues on the top of the barrel in (a); these surround the oxaloacetate/magnesium-binding site and extend through the cavity created by α-helices 2 and 3. The pink surface patches represent the charged residues on the top of the barrel. Figure 2. Sequence comparisons of citrate lyase β-subunit (CitE). The secondary structure of Mtb CitE highlights the multiple sequence alignment; red arrows depict the β-strands, the cream cylinders depict the α-helices, the green cylinder depicts α-helix 6a and the yellow cylinder depicts α-helix 6b; the α-helices and β-strand are numbered. Numbers above the sequence correspond to the residue numbers of Mtb CitE. Universally conserved residues are highlighted in blue; residues conserved only among genomes that lack the bacteria α and γ-subunits of citryl lyase are highlighted in pink; and residues involved in forming the trimer interface are highlighted in cyan. Abbreviation of the species names are as follows: M_tub; Mycobacterium tuberculosis, M_mus; Mus musculus, H_sap; Homo sapiens, D_rad; Deinococcus radiodurans, P_aer; Pseudomonas aeruginosa, B_par, Bordetella parapertussis, H_inf; Haemophilus influenzae, S_fle; Shigella flexneri, V_chl; Vibrio cholerae, S_pyo; Streptococcus pyogenes, K_pne ; Klebsiella pneumoniae, E_col; Escherichia coli, M_ext, Methylobacterium extorquens; and the last sequence in the alignment is the enzyme MclA, L-malyl-coenzymeA lyase from M_ext. It is noteworthy that the final six CitE sequences have known α and γ citrate lyase subunits of the bacterial citrate lyase complex. The sequence alignment was made with CLUSTALW and BoxShade. (b) Ribbon diagram of the trimer of Mtb CitE. The structure of CitE was determined by the multiwavelength anomalous diffraction (MAD) phasing method. Three data sets were collected for the SeMet protein in the R32 crystal form at wavelengths near the Se absorption edge at the synchrotron light source at Brookhaven National Laboratories on a charge-coupled device detector. Data were processed using DENZO and SCALEPACK,[4]^18 and MAD phasing proceeded by the standard methods of heavy-atom location (SHELXD [http://www.shelx.uni-ac.gwdg.de/SHELX/]), maximum likelihood phase refinement (ML-PHARE)[5]^26 and density modification [DM].[6]^19 Phase extension to 1.65 Å permitted automated model building for 70% of the protein with ARP/wARP.[7]^20 The remaining residues were traced with the program O.[8]^21 The model was then refined with the program CNS.[9]^22 The geometry of the structure was checked with ERRAT.[10]^23 The α-helices and β-strands are colored as in (a). The Figure was generated using PYMOL [http://www.pymol.sourceforge.net/]. (c) Illustration of the catalytic site of Mtb CitE. A cartoon of the catalytic site complexed with a magnesium ion and oxaloacetate. The ribbon diagram is colored the same as in (b), except the β-strands are colored wheat. The residues that form hydrogen bonds with oxaloacetate and the magnesium ion are shown in a stick representation; carbon, oxygen and nitrogen atoms are colored white, red and blue, respectively. The oxaloacetate is shown in stick form, its carbon and oxygen atoms are colored orange and red, the magnesium ion is shown as a yellow sphere and the water molecules are colored red. The hydrogen bonds are shown with broken black lines and the distance between the atoms is indicated. (d) Illustration of potential binding cavities for citryl-CoA. The illustration shows a ribbon diagram colored as in (c). The blue patches on the surface of the monomer are the conserved charged residues on the top of the barrel in (a); these surround the oxaloacetate/magnesium-binding site and extend through the cavity created by α-helices 2 and 3. The pink surface patches represent the charged residues on the top of the barrel.
 
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2007, 365, 275-283) copyright 2007.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20882276 B.E.Alber (2011).
Biotechnological potential of the ethylmalonyl-CoA pathway.
  Appl Microbiol Biotechnol, 89, 17-25.  
21083941 P.Sharma, B.Kumar, Y.Gupta, N.Singhal, V.M.Katoch, K.Venkatesan, and D.Bisht (2010).
Proteomic analysis of streptomycin resistant and sensitive clinical isolates of Mycobacterium tuberculosis.
  Proteome Sci, 8, 59.  
20047909 T.J.Erb, L.Frerichs-Revermann, G.Fuchs, and B.E.Alber (2010).
The apparent malate synthase activity of Rhodobacter sphaeroides is due to two paralogous enzymes, (3S)-Malyl-coenzyme A (CoA)/{beta}-methylmalyl-CoA lyase and (3S)- Malyl-CoA thioesterase.
  J Bacteriol, 192, 1249-1258.  
19098923 J.B.Bliska, and A.Casadevall (2009).
Intracellular pathogenic bacteria and fungi--a case of convergent evolution?
  Nat Rev Microbiol, 7, 165-171.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time.