EzCatDB: S00358

DB codeS00358
RLCP classification5.191.546000.23
CATH domainDomain 13.40.50.1820Catalytic domain
E.C.4.1.2.47
CSA1dwo,1qj4

CATH domainRelated DB codes (homologues)
3.40.50.1820S00544,S00344,S00517,S00525,S00526,S00720,S00723,S00724,S00725,S00919,S00057,S00374,S00345,S00347,S00348,S00346,S00350,S00352,S00353,S00355,S00356,D00189,D00210,D00539,T00253

Enzyme Name
Swiss-protKEGG

P52704P52705
Protein nameHydroxynitrilaseHydroxynitrilase(S)-hydroxynitrile lyase
(S)-cyanohydrin producing hydroxynitrile lyase
(S)-oxynitrilase
(S)-HbHNL
(S)-MeHNL
hydroxynitrile lyase
oxynitrilase
HbHNL
MeHNL
(S)-selective hydroxynitrile lyase
(S)-cyanohydrin carbonyl-lyase (cyanide forming)
SynonymsEC 4.1.2.37
EC 4.1.2.37) ((S)-acetone-cyanohydrin lyase
S)-acetone-cyanohydrin lyase) ((S)-hydroxynitrile lyase
Oxynitrilase
EC 4.1.2.37
EC 4.1.2.37) ((S)-acetone-cyanohydrin lyase
S)-acetone-cyanohydrin lyase) ((S)-hydroxynitrile lyase
Oxynitrilase

KEGG pathways
MAP codePathways
MAP00460Cyanoamino acid metabolism

Swiss-prot:Accession NumberP52704P52705
Entry nameHNL_HEVBRHNL_MANES
ActivityAn aliphatic (S)-hydroxynitrile = cyanide + an aliphatic aldehyde or ketone.,An aromatic (S)-hydroxynitrile = cyanide + an aromatic aldehyde.An aliphatic (S)-hydroxynitrile = cyanide + an aliphatic aldehyde or ketone.,An aromatic (S)-hydroxynitrile = cyanide + an aromatic aldehyde.
SubunitHomodimer.Homotrimer.
Subcellular location

Cofactor



SubstratesProducts
KEGG-idC18797C00177C00071C01450
Compound(S)-HydroxynitrilecyanideAldehydeKetone
Typecarbohydrateorganic ioncarbohydratecarbohydrate
1dwoAUnboundUnboundUnboundBound:ACN
1dwoBUnboundUnboundUnboundUnbound
1dwpAUnboundUnboundUnboundAnalogue:ACT
1dwpBUnboundUnboundUnboundAnalogue:ACT
1dwqAUnboundUnboundUnboundBound:ATO
1dwqBUnboundUnboundUnboundUnbound
1e89AUnboundUnboundUnboundUnbound
1e89BUnboundUnboundUnboundUnbound
1e8dABound:CNHUnboundUnboundUnbound
1e8dBBound:CNHUnboundUnboundUnbound
1eb8AUnboundUnboundUnboundUnbound
1eb8BUnboundUnboundUnboundUnbound
1eb9AUnboundUnboundBound:HBAUnbound
1eb9BUnboundUnboundBound:HBAUnbound
1qj4AUnboundUnboundUnboundUnbound
1yasAUnboundUnboundUnboundAnalogue:HIS
2yasAUnboundAnalogue:SCNUnboundUnbound
3yasAUnboundUnboundUnboundBound:ACN
4yasAUnboundUnboundUnboundUnbound
5yasAUnboundUnboundUnboundAnalogue:FAC
6yasAUnboundUnboundUnboundUnbound
7yasAUnboundUnboundUnboundUnbound

Active-site residues
resource
Swiss-prot;P52704, P52705 & literature [8], [11] & [12]
pdbCatalytic residuesModified residuescomment
1dwoATHR 11;SER 80;ASP 208;HIS 236


1dwoBTHR 11;SER 80;ASP 208;HIS 236


1dwpATHR 11;SER 80;ASP 208;HIS 236


1dwpBTHR 11;SER 80;ASP 208;HIS 236


1dwqATHR 11;SER 80;ASP 208;HIS 236
CSA 81(acetonylation)

1dwqBTHR 11;SER 80;ASP 208;HIS 236
CSA 81(acetonylation)

1e89ATHR 11;      ;ASP 208;HIS 236

mutant S80A
1e89BTHR 11;      ;ASP 208;HIS 236

mutant S80A
1e8dATHR 11;      ;ASP 208;HIS 236

mutant S80A
1e8dBTHR 11;      ;ASP 208;HIS 236

mutant S80A
1eb8ATHR 11;SER 80;ASP 208;HIS 236


1eb8BTHR 11;SER 80;ASP 208;HIS 236


1eb9ATHR 11;SER 80;ASP 208;HIS 236


1eb9BTHR 11;SER 80;ASP 208;HIS 236


1qj4ATHR 11;SER 80;ASP 207;HIS 235;LYS 236


1yasATHR 11;SER 80;ASP 207;HIS 235;LYS 236


2yasATHR 11;SER 80;ASP 207;HIS 235;LYS 236


3yasATHR 11;SER 80;ASP 207;HIS 235;LYS 236


4yasATHR 11;SER 80;ASP 207;HIS 235;LYS 236


5yasATHR 11;SER 80;ASP 207;HIS 235;LYS 236


6yasATHR 11;SER 80;ASP 207;HIS 235;LYS 236


7yasATHR 11;SER 80;ASP 207;HIS 235;LYS 236



References for Catalytic Mechanism
ReferencesSectionsNo. of steps in catalysis
[1]Fig.23
[2]Fig.8, p.813-8193
[3]Fig.4, p.258333
[5]Fig. 6, p.447-4483
[6]Scheme 3, p.186, p.1935
[7]Fig.2, p.997-9984
[8]Fig.6, p.19972
[11]Fig.5, p.1993
[12]Fig.2, p.1019-10203
[13]Fig.5, p.295-2981

references
[1]
PubMed ID8958122
JournalAnn N Y Acad Sci
Year1996
Volume799
Pages707-12
AuthorsHasslacher M, Schall M, Hayn M, Griengl H, Kohlwein SD, Schwab H
Title(S)-hydroxynitrile lyase from Hevea brasiliensis.
[2]
CommentsX-RAY CRYSTALLOGRAPHY (1.9 ANGSTROMS).
Medline ID96434327
PubMed ID8805565
JournalStructure
Year1996
Volume4
Pages811-22
AuthorsWagner UG, Hasslacher M, Griengl H, Schwab H, Kratky C
TitleMechanism of cyanogenesis: the crystal structure of hydroxynitrile lyase from Hevea brasiliensis.
Related PDB1yas
Related Swiss-protP52704
[3]
PubMed ID8824213
JournalJ Biol Chem
Year1996
Volume271
Pages25830-4
AuthorsWajant H, Pfizenmaier K
TitleIdentification of potential active-site residues in the hydroxynitrile lyase from Manihot esculenta by site-directed mutagenesis.
[4]
PubMed ID9325140
JournalProtein Expr Purif
Year1997
Volume11
Pages61-71
AuthorsHasslacher M, Schall M, Hayn M, Bona R, Rumbold K, Luckl J, Griengl H, Kohlwein SD, Schwab H
TitleHigh-level intracellular expression of hydroxynitrile lyase from the tropical rubber tree Hevea brasiliensis in microbial hosts.
[5]
PubMed ID9094745
JournalProteins
Year1997
Volume27
Pages438-49
AuthorsHasslacher M, Kratky C, Griengl H, Schwab H, Kohlwein SD
TitleHydroxynitrile lyase from Hevea brasiliensis: molecular characterization and mechanism of enzyme catalysis.
[6]
PubMed ID10407140
JournalBiochim Biophys Acta
Year1999
Volume1432
Pages185-93
AuthorsHanefeld U, Straathof AJ, Heijnen JJ
TitleStudy of the (S)-hydroxynitrile lyase from Hevea brasiliensis: mechanistic implications.
[7]
CommentsX-RAY CRYSTALLOGRAPHY (1.1 ANGSTROMS)
Medline ID99423043
PubMed ID10494852
JournalBiol Chem
Year1999
Volume380
Pages993-1000
AuthorsGruber K, Gugganig M, Wagner UG, Kratky C
TitleAtomic resolution crystal structure of hydroxynitrile lyase from Hevea brasiliensis.
Related PDB1qj4
Related Swiss-protP52704
[8]
CommentsX-RAY CRYSTALLOGRAPHY (1.72 ANGSTROMS).
Medline ID20014021
PubMed ID10548044
JournalProtein Sci
Year1999
Volume8
Pages1990-2000
AuthorsZuegg J, Gruber K, Gugganig M, Wagner UG, Kratky C
TitleThree-dimensional structures of enzyme-substrate complexes of the hydroxynitrile lyase from Hevea brasiliensis.
Related PDB2yas,3yas,4yas,5yas,6yas,7yas
Related Swiss-protP52704
[9]
PubMed ID11102786
JournalCurr Opin Biotechnol
Year2000
Volume11
Pages532-9
AuthorsEffenberger F, Forster S, Wajant H
TitleHydroxynitrile lyases in stereoselective catalysis.
[10]
PubMed ID10679367
JournalCurr Opin Chem Biol
Year2000
Volume4
Pages103-9
AuthorsJohnson DV, Zabelinskaja-Mackova AA, Griengl H
TitleOxynitrilases for asymmetric C-C bond formation.
[11]
CommentsX-ray crystallography
PubMed ID11173464
JournalActa Crystallogr D Biol Crystallogr
Year2001
Volume57
Pages194-200
AuthorsLauble H, Forster S, Miehlich B, Wajant H, Effenberger F
TitleStructure of hydroxynitrile lyase from Manihot esculenta in complex with substrates acetone and chloroacetone: implications for the mechanism of cyanogenesis.
Related PDB1dwo,1dwp,1dwq
[12]
PubMed ID11316882
JournalProtein Sci
Year2001
Volume10
Pages1015-22
AuthorsLauble H, Miehlich B, Forster S, Wajant H, Effenberger F
TitleMechanistic aspects of cyanogenesis from active-site mutant Ser80Ala of hydroxynitrile lyase from Manihot esculenta in complex with acetone cyanohydrin.
Related PDB1e89,1e8d
[13]
PubMed ID11790839
JournalProtein Sci
Year2002
Volume11
Pages292-300
AuthorsDreveny I, Kratky C, Gruber K
TitleThe active site of hydroxynitrile lyase from Prunus amygdalus: modeling studies provide new insights into the mechanism of cyanogenesis.
[14]
PubMed ID11742123
JournalProtein Sci
Year2002
Volume11
Pages65-71
AuthorsLauble H, Miehlich B, Forster S, Kobler C, Wajant H, Effenberger F
TitleStructure determinants of substrate specificity of hydroxynitrile lyase from Manihot esculenta.
Related PDB1eb8,1eb9
[15]
PubMed ID12889812
JournalBiotechnol Lett
Year2003
Volume25
Pages1041-7
AuthorsYan G, Cheng S, Zhao G, Wu S, Liu Y, Sun W
TitleA single residual replacement improves the folding and stability of recombinant cassava hydroxynitrile lyase in E. coil.

comments
This enzyme belongs to the alpha/beta hydrolase superfamily, which has a catalytic triad (Ser/Asp/His).
The earlier literature ([1], [2] & [5]) proposed a catalytic mechanism, in which Ser80 acts as a nucleophilic residue to form a tetrahedral intermediate with substrate, as that in serine proteases.
However, more recent papers ([3], [6], [7], [8], [11] & [12]) proposed an alternative mechanism, in which catalytic residues act as acid/base or stabilizers.
This enzyme catalyzes a reversible reaction. In the forward direction, cyanohydrin will be degradaded into acetone and cyanide, accompanied by elimination and double-bond formation, whilst cyanohydrin will be synthesized from acetone and cyanide, by addition to the double-bond, in the reverse direction.
The forward reaction (elimination and double-bond formation reaction) proceeds as follows (see ([3], [6], [7], [8], [11] & [12]):
(1) Ser80, modulated by His235, acts as a general base, to abstract a proton from cyanohydrin OH-group, leading to the elimination of the cyanide (or nitrile). During this reaction, Thr11 stabilizes the oxygen atom of the OH-group, whilst His235 and Lys236 (PDB; 1qj4; enzyme from Havea brasiliensis) stabilize the eliminated group, cyanide (or nitrile).
(2) His235 acts as a general acid, to donate a proton to the eliminated group, the cyanide ion.
In the enzyme from Manihot esculenta, however, the corresponding lysine residue, Lys237, does not seem to be involved in the stabilization, according to the literature [11] & [12].
The reverse reaction (addition to double-bond) adopts an analogous mechanism, using the same catalytic residues.

createdupdated
2004-04-042012-10-03
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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