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PDBsum entry 1bli

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protein metals links
Hydrolase PDB id
1bli
Jmol
Contents
Protein chain
481 a.a. *
Metals
_NA
_CA ×3
Waters ×221
* Residue conservation analysis
PDB id:
1bli
Name: Hydrolase
Title: Bacillus licheniformis alpha-amylase
Structure: Alpha-amylase. Chain: a. Synonym: glucanotransferase, alpha-1,4-glucan-4- glucanohydrolase. Engineered: yes. Mutation: yes. Other_details: protein produced by novo nordisk a/s, denmark
Source: Bacillus licheniformis. Organism_taxid: 1402. Cell_line: 293. Atcc: atcc 6598. Collection: atcc 6598. Gene: amyl. Expressed in: bacillus subtilis. Expression_system_taxid: 1423.
Resolution:
1.90Å     R-factor:   0.154     R-free:   0.185
Authors: M.Machius,N.Declerck,R.Huber,G.Wiegand
Key ref:
M.Machius et al. (1998). Activation of Bacillus licheniformis alpha-amylase through a disorder-->order transition of the substrate-binding site mediated by a calcium-sodium-calcium metal triad. Structure, 6, 281-292. PubMed id: 9551551 DOI: 10.1016/S0969-2126(98)00032-X
Date:
07-Jan-98     Release date:   23-Mar-99    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P06278  (AMY_BACLI) -  Alpha-amylase
Seq:
Struc:
512 a.a.
481 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 7 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.3.2.1.1  - Alpha-amylase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Endohydrolysis of 1,4-alpha-glucosidic linkages in oligosaccharides and polysaccharides.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     metabolic process   2 terms 
  Biochemical function     catalytic activity     8 terms  

 

 
DOI no: 10.1016/S0969-2126(98)00032-X Structure 6:281-292 (1998)
PubMed id: 9551551  
 
 
Activation of Bacillus licheniformis alpha-amylase through a disorder-->order transition of the substrate-binding site mediated by a calcium-sodium-calcium metal triad.
M.Machius, N.Declerck, R.Huber, G.Wiegand.
 
  ABSTRACT  
 
BACKGROUND: The structural basis as to how metals regulate the functional state of a protein by altering or stabilizing its conformation has been characterized in relatively few cases because the metal-free form of the protein is often partially disordered and unsuitable for crystallographic analysis. This is not the case, however, for Bacillus licheniformis alpha-amylase (BLA) for which the structure of the metal-free form is available. BLA is a hyperthermostable enzyme which is widely used in biotechnology, for example in the breakdown of starch or as a component of detergents. The determination of the structure of BLA in the metal-containing form, together with comparisons to the apo enzyme, will help us to understand the way in which metal ions can regulate enzyme activity. RESULTS: We report here the crystal structure of native, metal-containing BLA. The structure shows that the calcium-binding site which is conserved in all alpha-amylases forms part of an unprecedented linear triadic metal array, with two calcium ions flanking a central sodium ion. A region around the metal triad comprising 21 residues exhibits a conformational change involving a helix unwinding and a disorder-->order transition compared to the structure of metal-free BLA. Another calcium ion, not previously observed in alpha-amylases, is located at the interface between domains A and C. CONCLUSIONS: We present a structural description of a major conformational rearrangement mediated by metal ions. The metal induced disorder-->order transition observed in BLA leads to the formation of the extended substrate-binding site and explains on a structural level the calcium dependency of alpha-amylases. Sequence comparisons indicate that the unique Ca-Na-Ca metal triad and the additional calcium ion located between domains A and C might be found exclusively in bacterial alpha-amylases which show increased thermostability. The information presented here may help in the rational design of mutants with enhanced performance in biotechnological applications.
 
  Selected figure(s)  
 
Figure 6.
Figure 6. Stereo view CPK representation of BLA with the carbohydrate inhibitor V-1532 from the structure of the PPA-V-1532 complex [23] superimposed. The region which is ordered only in metal-containing BLA is shown in red; calcium ions are in cyan and sodium is in yellow. Residues in domain A of BLA which are homologous to sugar-binding residues in domain A of PPA are shown in green and the pseudo-oligosaccharide inhibitor V-1532 from the PPA-V-1532 complex structure is in blue stick form (see text for details).
 
  The above figure is reprinted by permission from Cell Press: Structure (1998, 6, 281-292) copyright 1998.  
  Figure was selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19763902 O.Prakash, and N.Jaiswal (2010).
alpha-Amylase: an ideal representative of thermostable enzymes.
  Appl Biochem Biotechnol, 160, 2401-2414.  
21092126 S.Kumar, N.Singh, B.Mishra, D.Dube, M.Sinha, S.B.Singh, S.Dey, P.Kaur, S.Sharma, and T.P.Singh (2010).
Modulation of inhibitory activity of xylanase-α-amylase inhibitor protein (XAIP): binding studies and crystal structure determination of XAIP-II from Scadoxus multiflorus at 1.2 Å resolution.
  BMC Struct Biol, 10, 41.
PDB code: 3mu7
18951544 B.Khemakhem, M.B.Ali, N.Aghajari, M.Juy, R.Haser, and S.Bejar (2009).
Engineering of the alpha-amylase from Geobacillus stearothermophilus US100 for detergent incorporation.
  Biotechnol Bioeng, 102, 380-389.  
18451358 B.F.Shaw, G.F.Schneider, B.Bilgiçer, G.K.Kaufman, J.M.Neveu, W.S.Lane, J.P.Whitelegge, and G.M.Whitesides (2008).
Lysine acetylation can generate highly charged enzymes with increased resistance toward irreversible inactivation.
  Protein Sci, 17, 1446-1455.  
18473149 Y.Xu, M.Yang, J.Sun, J.Qian, D.Zhang, Y.Sun, L.Ma, and C.Zhu (2008).
Glycogen branching enzyme: a novel deltamethrin resistance-associated gene from Culex pipiens pallens.
  Parasitol Res, 103, 449-458.  
17598074 R.Priyadharshini, and P.Gunasekaran (2007).
Site-directed mutagenesis of the calcium-binding site of alpha-amylase of Bacillus licheniformis.
  Biotechnol Lett, 29, 1493-1499.  
17310272 S.Srimathi, G.Jayaraman, G.Feller, B.Danielsson, and P.R.Narayanan (2007).
Intrinsic halotolerance of the psychrophilic alpha-amylase from Pseudoalteromonas haloplanktis.
  Extremophiles, 11, 505-515.  
17154418 T.Shirai, K.Igarashi, T.Ozawa, H.Hagihara, T.Kobayashi, K.Ozaki, and S.Ito (2007).
Ancestral sequence evolutionary trace and crystal structure analyses of alkaline alpha-amylase from Bacillus sp. KSM-1378 to clarify the alkaline adaptation process of proteins.
  Proteins, 66, 600-610.
PDB code: 2die
16267046 E.Di Cera (2006).
A structural perspective on enzymes activated by monovalent cations.
  J Biol Chem, 281, 1305-1308.  
16710633 K.Hirasawa, K.Uchimura, M.Kashiwa, W.D.Grant, S.Ito, T.Kobayashi, and K.Horikoshi (2006).
Salt-activated endoglucanase of a strain of alkaliphilic Bacillus agaradhaerens.
  Antonie Van Leeuwenhoek, 89, 211-219.  
16452622 R.Kanai, K.Haga, T.Akiba, K.Yamane, and K.Harata (2006).
Role of Trp140 at subsite -6 on the maltohexaose production of maltohexaose-producing amylase from alkalophilic Bacillus sp.707.
  Protein Sci, 15, 468-477.
PDB codes: 2d3l 2d3n
15610414 M.Shahhoseini, A.A.Ziaee, A.A.Pourbabai, N.Ghaemi, and N.Declerck (2005).
A natural variant of Bacillus licheniformis alpha-amylase isolated from flour mill wastewaters sheds light on the origin of high thermostability.
  J Appl Microbiol, 98, 24-32.  
15722449 R.Maurus, A.Begum, H.H.Kuo, A.Racaza, S.Numao, C.Andersen, J.W.Tams, J.Vind, C.M.Overall, S.G.Withers, and G.D.Brayer (2005).
Structural and mechanistic studies of chloride induced activation of human pancreatic alpha-amylase.
  Protein Sci, 14, 743-755.
PDB codes: 1xgz 1xh0 1xh1 1xh2
14760745 A.Linden, and M.Wilmanns (2004).
Adaptation of class-13 alpha-amylases to diverse living conditions.
  Chembiochem, 5, 231-239.  
14660599 K.S.Bak-Jensen, G.André, T.E.Gottschalk, G.Paës, V.Tran, and B.Svensson (2004).
Tyrosine 105 and threonine 212 at outermost substrate binding subsites -6 and +4 control substrate specificity, oligosaccharide cleavage patterns, and multiple binding modes of barley alpha-amylase 1.
  J Biol Chem, 279, 10093-10102.  
12482867 A.Linden, O.Mayans, W.Meyer-Klaucke, G.Antranikian, and M.Wilmanns (2003).
Differential regulation of a hyperthermophilic alpha-amylase with a novel (Ca,Zn) two-metal center by zinc.
  J Biol Chem, 278, 9875-9884.
PDB codes: 1mwo 1mxd 1mxg
  16233519 A.Tanaka, and E.Hoshino (2003).
Secondary calcium-binding parameter of Bacillus amyloliquefaciens alpha-amylase obtained from inhibition kinetics.
  J Biosci Bioeng, 96, 262-267.  
12930989 J.E.Nielsen, and J.A.McCammon (2003).
Calculating pKa values in enzyme active sites.
  Protein Sci, 12, 1894-1901.  
12540849 M.Machius, N.Declerck, R.Huber, and G.Wiegand (2003).
Kinetic stabilization of Bacillus licheniformis alpha-amylase through introduction of hydrophobic residues at the surface.
  J Biol Chem, 278, 11546-11553.
PDB code: 1ob0
12676725 R.J.Shiau, H.C.Hung, and C.L.Jeang (2003).
Improving the thermostability of raw-starch-digesting amylase from a Cytophaga sp. by site-directed mutagenesis.
  Appl Environ Microbiol, 69, 2383-2385.  
12719434 T.Nonaka, M.Fujihashi, A.Kita, H.Hagihara, K.Ozaki, S.Ito, and K.Miki (2003).
Crystal structure of calcium-free alpha-amylase from Bacillus sp. strain KSM-K38 (AmyK38) and its sodium ion binding sites.
  J Biol Chem, 278, 24818-24824.
PDB codes: 1ud2 1ud3 1ud4 1ud5 1ud6 1ud8
12906828 X.Robert, R.Haser, T.E.Gottschalk, F.Ratajczak, H.Driguez, B.Svensson, and N.Aghajari (2003).
The structure of barley alpha-amylase isozyme 1 reveals a novel role of domain C in substrate recognition and binding: a pair of sugar tongs.
  Structure, 11, 973-984.
PDB codes: 1ht6 1p6w
12021442 N.Aghajari, G.Feller, C.Gerday, and R.Haser (2002).
Structural basis of alpha-amylase activation by chloride.
  Protein Sci, 11, 1435-1441.
PDB codes: 1jd7 1jd9 1l0p
11257505 E.A.MacGregor, S.Janecek, and B.Svensson (2001).
Relationship of sequence and structure to specificity in the alpha-amylase family of enzymes.
  Biochim Biophys Acta, 1546, 1.  
11453991 H.Hagihara, Y.Hayashi, K.Endo, K.Igarashi, T.Ozawa, S.Kawai, K.Ozaki, and S.Ito (2001).
Deduced amino-acid sequence of a calcium-free alpha-amylase from a strain of Bacillus: implications from molecular modeling of high oxidation stability and chelator resistance of the enzyme.
  Eur J Biochem, 268, 3974-3982.  
11418770 S.Korolev, I.Dementieva, R.Sanishvili, W.Minor, Z.Otwinowski, and A.Joachimiak (2001).
Using surface-bound rubidium ions for protein phasing.
  Acta Crystallogr D Biol Crystallogr, 57, 1008-1012.  
10924103 A.M.Brzozowski, D.M.Lawson, J.P.Turkenburg, H.Bisgaard-Frantzen, A.Svendsen, T.V.Borchert, Z.Dauter, K.S.Wilson, and G.J.Davies (2000).
Structural analysis of a chimeric bacterial alpha-amylase. High-resolution analysis of native and ligand complexes.
  Biochemistry, 39, 9099-9107.
PDB codes: 1e3x 1e3z 1e40 1e43
10666605 D.Suvd, K.Takase, Z.Fujimoto, M.Matsumura, and H.Mizuno (2000).
Purification, crystallization and preliminary X-ray crystallographic study of alpha-amylase from Bacillus stearothermophilus.
  Acta Crystallogr D Biol Crystallogr, 56, 200-202.  
11150610 J.E.Nielsen, and T.V.Borchert (2000).
Protein engineering of bacterial alpha-amylases.
  Biochim Biophys Acta, 1543, 253-274.  
10969023 J.Fitter, and J.Heberle (2000).
Structural equilibrium fluctuations in mesophilic and thermophilic alpha-amylase.
  Biophys J, 79, 1629-1636.  
10672010 K.W.Rodenburg, F.Vallée, N.Juge, N.Aghajari, X.Guo, R.Haser, and B.Svensson (2000).
Specific inhibition of barley alpha-amylase 2 by barley alpha-amylase/subtilisin inhibitor depends on charge interactions and can be conferred to isozyme 1 by mutation.
  Eur J Biochem, 267, 1019-1029.  
10449318 A.Shaw, R.Bott, and A.G.Day (1999).
Protein engineering of alpha-amylase for low pH performance.
  Curr Opin Biotechnol, 10, 349-352.  
10491128 J.E.Nielsen, L.Beier, D.Otzen, T.V.Borchert, H.B.Frantzen, K.V.Andersen, and A.Svendsen (1999).
Electrostatics in the active site of an alpha-amylase.
  Eur J Biochem, 264, 816-824.  
10540739 K.Igarashi, T.Ozawa, K.Ikawakitayama, Y.Hayashi, H.Araki, K.Endo, H.Hagihara, K.Ozaki, S.Kawai, and S.Ito (1999).
Thermostabilization by proline substitution in an alkaline, liquefying alpha-amylase from Bacillus sp. strain KSM-1378.
  Biosci Biotechnol Biochem, 63, 1535-1540.  
10099546 L.M.Marchal, J.Jonkers, G.T.Franke, C.D.de Gooijer, and J.Tramper (1999).
The effect of process conditions on the alpha-amylolytic hydrolysis of amylopectin potato starch: An experimental design approach.
  Biotechnol Bioeng, 62, 348-357.  
9862804 N.Aghajari, G.Feller, C.Gerday, and R.Haser (1998).
Structures of the psychrophilic Alteromonas haloplanctis alpha-amylase give insights into cold adaptation at a molecular level.
  Structure, 6, 1503-1516.
PDB code: 1b0i
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. Where a reference describes a PDB structure, the PDB code is shown on the right.