EzCatDB: S00437
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DB codeS00437
RLCP classification1.13.30185.55 : Hydrolysis
CATH domainDomain 13.60.60.10 : Penicillin V Acylase; Chain ACatalytic domain
E.C.3.5.1.11
MACiEM0241


Enzyme Name
UniProtKBKEGG

P12256
Protein namePenicillin acylasepenicillin amidase
penicillin acylase
benzylpenicillin acylase
novozym 217
semacylase
alpha-acylamino-beta-lactam acylhydrolase
ampicillin acylase
SynonymsEC 3.5.1.11
Penicillin V amidase
PVA
MEROPSC59.001 (Cysteine)
PfamPF02275 (CBAH)
[Graphical view]

KEGG pathways
MAP codePathways
MAP00311Penicillin and cephalosporin biosynthesis

UniProtKB:Accession NumberP12256
Entry namePAC_BACSH
ActivityPenicillin + H(2)O = a carboxylate + 6- aminopenicillanate.
SubunitHomotetramer.
Subcellular location
Cofactor

Compound table: links to PDB-related databases & PoSSuM

SubstratesProducts
KEGG-idC00395C00001C00060C02954
CompoundPenicillinH2OCarboxylate6-Aminopenicillanate
Typecarboxyl group,peptide/protein,sulfide groupH2Ocarboxyl groupamide group,amine group,carboxyl group,sulfide group
ChEBI
15377

16705
57869
PubChem
962
22247451

7057887
11082
            
2pvaAUnbound UnboundUnbound
2pvaBUnbound UnboundUnbound
2pvaCUnbound UnboundUnbound
2pvaDUnbound UnboundUnbound
3pvaAUnbound UnboundUnbound
3pvaBUnbound UnboundUnbound
3pvaCUnbound UnboundUnbound
3pvaDUnbound UnboundUnbound
3pvaEUnbound UnboundUnbound
3pvaFUnbound UnboundUnbound
3pvaGUnbound UnboundUnbound
3pvaHUnbound UnboundUnbound

Active-site residues
resource
literature [1] & [4]
pdbCatalytic residuesModified residuesMain-chain involved in catalysiscomment
            
2pvaA     ;ASP 20;ASN 175;ARG 228
OCS 1
TYR 82
OCS,Cysteinesulfonic acid
2pvaB     ;ASP 20;ASN 175;ARG 228
OCS 1
TYR 82
OCS,Cysteinesulfonic acid
2pvaC     ;ASP 20;ASN 175;ARG 228
OCS 1
TYR 82
OCS,Cysteinesulfonic acid
2pvaD     ;ASP 20;ASN 175;ARG 228
OCS 1
TYR 82
OCS,Cysteinesulfonic acid
3pvaACYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 
3pvaBCYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 
3pvaCCYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 
3pvaDCYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 
3pvaECYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 
3pvaFCYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 
3pvaGCYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 
3pvaHCYS 1;ASP 20;ASN 175;ARG 228
     
TYR 82
 

References for Catalytic Mechanism
ReferencesSectionsNo. of steps in catalysis
[1]Fig.35
[3]p.239
[4]p.414
[6]Fig.23
[7]p.146-147

references
[1]
CommentsPenicillin G acylase (homologous enzyme)
PubMed ID7816145
JournalNature
Year1995
Volume373
Pages264-8
AuthorsDuggleby HJ, Tolley SP, Hill CP, Dodson EJ, Dodson G, Moody PC
TitlePenicillin acylase has a single-amino-acid catalytic centre.
[2]
CommentsPenicillin G acylase (homologous enzyme)
PubMed ID7477383
JournalNature
Year1995
Volume378
Pages416-9
AuthorsBrannigan JA, Dodson G, Duggleby HJ, Moody PC, Smith JL, Tomchick DR, Murzin AG
TitleA protein catalytic framework with an N-terminal nucleophile is capable of self-activation.
[3]
CommentsPenicillin G acylase (homologous enzyme)
PubMed ID9931321
JournalBiochem J
Year1999
Volume338
Pages235-9
AuthorsMorillas M, Goble ML, Virden R
TitleThe kinetics of acylation and deacylation of penicillin acylase from Escherichia coli ATCC 11105: evidence for lowered pKa values of groups near the catalytic centre.
[4]
CommentsX-ray crystallography
PubMed ID10331865
JournalNat Struct Biol
Year1999
Volume6
Pages414-416
AuthorsSuresh CG, Pundle AV, SivaRaman H, Rao KN, Brannigan JA, McVey CE, Verma CS, Dauter Z, Dodson EJ, Dodson GG
TitlePenicillin V acylase crystal structure reveals new Ntn-hydrolase family members.
Related PDB2pva,3pva
[5]
CommentsPenicillin G acylase (homologous enzyme)
PubMed ID10993730
JournalJ Mol Biol
Year2000
Volume302
Pages887-98
AuthorsHewitt L, Kasche V, Lummer K, Lewis RJ, Murshudov GN, Verma CS, Dodson GG, Wilson KS
TitleStructure of a slow processing precursor penicillin acylase from Escherichia coli reveals the linker peptide blocking the active-site cleft.
[6]
CommentsX-ray crystallography, catalysis; Penicillin G acylase (homologous enzyme)
PubMed ID11239085
JournalProtein Eng
Year2000
Volume13
Pages857-863
AuthorsAlkema WB, Hensgens CM, Kroezinga EH, de Vries E, Floris R, van der Laan JM, Dijkstra BW, Janssen DB
TitleCharacterization of the beta-lactam binding site of penicillin acylase of Escherichia coli by structural and site-directed mutagenesis studies.
[7]
CommentsX-ray crystallography (1.3 Angstroms), catalysis; Penicillin G acylase (homologous enzyme)
PubMed ID11601852
JournalJ Mol Biol
Year2001
Volume313
Pages139-150
AuthorsMcVey CE, Walsh MA, Dodson GG, Wilson KS, Brannigan JA
TitleCrystal structures of penicillin acylase enzyme-substrate complexes: structural insights into the catalytic mechanism.

comments
This enzyme belongs to N-terminal nucleophile (Ntn) hydrolase family, of which sidechain of the N-terminal residue acts as nucleophile, assisted by its own alpha-amino group.
The paper [1] proposed the catalytic mechanism of the homologous enzyme, penicillin G acylase, whose nucleophilic residue is serine instead of cysteine.
(1) The alpha-amino group enhances the nucleophilicity of the sidechain of the N-terminal residue, through the bridging water between them.
(2) The sidechain of Cys1 makes a nucleophilic attack on the acyl carbon of penicillin, forming an oxyanion tetrahedral, stabilized by the oxyanion hole composed of mainchain amides and sidechain of Asn175. Here, the alpha-amino group protonates the leaving amine group through a water.
(3) This tetrahedral intermediate will then collapse to form an acyl enzyme and release the free 6-aminopenicillanic acid.
(4) The acyl enzyme will be attacked by water to form a second tetrahedral intermediate, stabilized by the same oxyanion hole, which can in turn collapse to release the free phenylacetic acid.
In contrast, the literature [7] suggested that the catalytic mechanism proceeds via direct nucleophilic attack of the nucleophilic residue on the scissile amide and not as via the bridging water molecule acting as a "virtual" base.
According to the paper [3], the residue corresponding to Arg228 in its homologue, penicillin G acylase, seems to be important for catalysis, orienting the N-terminal catalytic residue and contributing to a decrease in the pKa of alpha-amino group.
The literature on this enzyme [4] reported that the oxyanion hole consists of the sidechain of Asn175 and NH of Tyr82, whilst Arg228, Asp20 and Asn175 are critical for positioning the lone pair of the unprotonated N-terminal alpha-amino group. This might lead to the decrease in the pKa of the alpha-amino group.

createdupdated
2002-09-042009-02-26


Copyright: Nozomi Nagano, JST & CBRC-AIST
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Funded by BIRD/Japan Science and Technology Corporation (JST) (October 2007 - September 2010)
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