EzCatDB: S00394

DB codeS00394
RLCP classification1.13.11100.261
CATH domainDomain 13.40.390.10Catalytic domain
E.C.3.4.24.21
CSA1ast

CATH domainRelated DB codes (homologues)
3.40.390.10S00395,S00397,S00398,S00399,D00232,D00236,M00101

Enzyme Name
Swiss-protKEGG

P07584
Protein nameAstacinastacin
Astacus proteinase
crayfish small-molecule proteinase
SynonymsEC 3.4.24.21
Crayfish small molecule proteinase


Swiss-prot:Accession NumberP07584
Entry nameASTA_ASTFL
ActivityHydrolysis of peptide bonds in substrates containing five or more amino acids, preferentially with Ala in P1'', and Pro in P2''.
SubunitMonomer.
Subcellular location
CofactorBinds 1 zinc ion per subunit.


CofactorsSubstratesProductsintermediates
KEGG-idC00038C00017C00012C00001C00017C00012
CompoundZincProteinPeptideH2OProteinPeptide
Typeheavy metalpeptide/proteinpeptide/proteinH2Opeptide/proteinpeptide/protein
1astABound:_ZNUnboundUnbound
UnboundUnboundUnbound
1iaaAAnalogue:_CUUnboundUnbound
UnboundUnboundUnbound
1iabAAnalogue:_COUnboundUnbound
UnboundUnboundUnbound
1iacAAnalogue:_HGUnboundUnbound
UnboundUnboundUnbound
1iadAUnboundUnboundUnbound
UnboundUnboundUnbound
1iaeAAnalogue:_NIUnboundUnbound
UnboundUnboundUnbound
1qjiABound:_ZNUnboundUnbound
UnboundUnboundTransition-state-analogue:PKF
1qjjABound:_ZNUnboundUnbound
UnboundAnalogue:PRO-LEU-GLY-HOAUnbound

Active-site residues
resource
Swiss-prot;P07584
pdbCatalytic residuesCofactor-binding residues
1astAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)
1iaaAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)
1iabAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)
1iacAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)
1iadAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)
1iaeAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)
1qjiAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)
1qjjAGLU 93;TYR 149
HIS 92;HIS 96;HIS 102;TYR 149(Zinc binding)

References for Catalytic Mechanism
ReferencesSectionsNo. of steps in catalysis
[6]p.17113-17117
[7]p.321-325
[9]Fig.3, p.673-6743
[10]Scheme 3, p.4955-4958
[11]p.227

references
[1]
CommentsX-ray crystallography
PubMed ID1319561
JournalNature
Year1992
Volume358
Pages164-7
AuthorsBode W, Gomis-Ruth FX, Huber R, Zwilling R, Stocker W
TitleStructure of astacin and implications for activation of astacins and zinc-ligation of collagenases.
Related PDB1ast
Related Swiss-protP07584
[2]
CommentsX-ray crystallography (1.8 Angstroms)
PubMed ID8445658
JournalJ Mol Biol
Year1993
Volume229
Pages945-68
AuthorsGomis-Ruth FX, Stocker W, Huber R, Zwilling R, Bode W
TitleRefined 1.8 A X-ray crystal structure of astacin, a zinc-endopeptidase from the crayfish Astacus astacus L. Structure determination, refinement, molecular structure and comparison with thermolysin.
Related PDB1iac,1iad
Related Swiss-protP07584
[3]
PubMed ID8248170
JournalProc Natl Acad Sci U S A
Year1993
Volume90
Pages10783-7
AuthorsWolfsberg TG, Bazan JF, Blobel CP, Myles DG, Primakoff P, White JM
TitleThe precursor region of a protein active in sperm-egg fusion contains a metalloprotease and a disintegrin domain: structural, functional, and evolutionary implications.
[4]
PubMed ID8262184
JournalFEBS Lett
Year1993
Volume335
Pages361-6
AuthorsCorbeil D, Milhiet PE, Simon V, Ingram J, Kenny AJ, Boileau G, Crine P
TitleRat endopeptidase-24.18 alpha subunit is secreted into the culture medium as a zymogen when expressed by COS-1 cells.
[5]
PubMed ID8198548
JournalBiochem J
Year1994
Volume300
Pages37-43
AuthorsMilhiet PE, Corbeil D, Simon V, Kenny AJ, Crine P, Boileau G
TitleExpression of rat endopeptidase-24.18 in COS-1 cells: membrane topology and activity.
[6]
CommentsX-ray crystallography
PubMed ID8006015
JournalJ Biol Chem
Year1994
Volume269
Pages17111-7
AuthorsGomis-Ruth FX, Grams F, Yiallouros I, Nar H, Kusthardt U, Zwilling R, Bode W, Stocker W
TitleCrystal structures, spectroscopic features, and catalytic properties of cobalt(II), copper(II), nickel(II), and mercury(II) derivatives of the zinc endopeptidase astacin. A correlation of structure and proteolytic activity.
Related PDB1iaa,1iab,1iae
[7]
PubMed ID7674929
JournalMethods Enzymol
Year1995
Volume248
Pages305-25
AuthorsStocker W, Zwilling R
TitleAstacin.
[8]
PubMed ID8740360
JournalStructure
Year1996
Volume4
Pages375-86
AuthorsDhanaraj V, Ye QZ, Johnson LL, Hupe DJ, Ortwine DF, Dunbar JB Jr, Rubin JR, Pavlovsky A, Humblet C, Blundell TL
TitleX-ray structure of a hydroxamate inhibitor complex of stromelysin catalytic domain and its comparison with members of the zinc metalloproteinase superfamily.
[9]
PubMed ID8756323
JournalNat Struct Biol
Year1996
Volume3
Pages671-5
AuthorsGrams F, Dive V, Yiotakis A, Yiallouros I, Vassiliou S, Zwilling R, Bode W, Stocker W
TitleStructure of astacin with a transition-state analogue inhibitor.
Related PDB1qji,1qjj
[10]
PubMed ID9125517
JournalBiochemistry
Year1997
Volume36
Pages4949-58
AuthorsMock WL, Yao J
TitleKinetic characterization of the serralysins: a divergent catalytic mechanism pertaining to astacin-type metalloproteases.
[11]
PubMed ID11078883
JournalFEBS Lett
Year2000
Volume484
Pages224-8
AuthorsYiallouros I, Grosse Berkhoff E, Stocker W
TitleThe roles of Glu93 and Tyr149 in astacin-like zinc peptidases.
[12]
PubMed ID12441103
JournalJ Mol Biol
Year2002
Volume324
Pages237-46
AuthorsYiallouros I, Kappelhoff R, Schilling O, Wegmann F, Helms MW, Auge A, Brachtendorf G, Berkhoff EG, Beermann B, Hinz HJ, Konig S, Peter-Katalinic J, Stocker W
TitleActivation mechanism of pro-astacin: role of the pro-peptide, tryptic and autoproteolytic cleavage and importance of precise amino-terminal processing.

comments
This enzyme belongs to the peptidase family-M12A.
Although an alternative machanism has been reported [10], in which Tyr149, instead of Glu93, functions as a general base to deprotonate a solvent water molecule, other experimental study such as X-ray crystallography and mutagenesis ([9] & [11]) confirmed that Glu93 and Tyr149 act as a general base and a stabilizer of the transition state, respectively.
In the free enzyme, the catalytic zinc is liganded by three imidazol nitrogen atoms from the histidines (His92, His96 & His102) and a oxygen atom of solvent water molecule, and another oxygen atom from the sidechain of Tyr149, forming a trigonal-bipyramidal coordination sphere (see [2], [6] & [7]). Upon substrate binding, some rearrangement of the Tyr149 residue seems to occur, suggesting this is a 'tyrosine switch', according to the literature [9].
Thus, the catalytic mechanism proceeds as follows (see [9], [11]):
(1) Glu93 activates the catalytic water, which in turn makes a nucleophilic attack on the carbonyl carbon atom.
(2) Tyr149 acts as an electrophile, to bind and to stabilize the tetrahedral carboxyanion of the transition state.
(3) The protonated carboxylate of Glu93 then protonates the leaving amide group.

createdupdated
2002-09-272009-02-26
Copyright: Nozomi Nagano, JST & CBRC-AIST
Funded by PRESTO/Japan Science and Technology Corporation (JST) (December 2001 - November 2004)
Funded by Grant-in-Aid for Publication of Scientific Research Results/Japan Society for the Promotion of Science (JSPS) (April 2005 - March 2006)
Funded by Grant-in-Aid for Scientific Research (B)/Japan Society for the Promotion of Science (JSPS) (April 2005 - March 2008)
Funded by BIRD/Japan Science and Technology Corporation (JST) (September 2005 - September 2010)
Funded by BIRD/Japan Science and Technology Corporation (JST) (October 2007 - September 2010)
Funded by Grant-in-Aid for Publication of Scientific Research Results/Japan Society for the Promotion of Science (JSPS) (April 2011 - March 2012)

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