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Catalytic Site Atlas Version 2.2.12
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CSA entry for 1m21
Original Entry
Title:
Hydrolase
Compound:
Peptide amidase
Mutant:
No
UniProt/Swiss-Prot:
Q8RJN5-Q8RJN5
EC Class:
3.5.1.-
Other CSA Entries:
Overview of all sites for 1m21
Homologues of 1m21
Entries for UniProt/Swiss-Prot: Q8RJN5
Entries for EC: 3.5.1.-
Other Databases:
PDB entry: 1m21
PDBsum entry: 1m21
UniProt/Swiss-Prot: Q8RJN5
IntEnz entry: 3.5.1.-
Literature Report:
Introduction:
Peptide amidase (PAM) from Strenotrophomonas maltophilia (a gram-negative bacterium) catalyses the hydrolysis of the C-terminal amide bond in peptide amides. It is very regio-selective, and those terminal bonds in amino acid side chains are not hydrolysed. PAM belongs to the amidase signature (AS) family, most of which display hydrolase activity. The natural function of periplasmatic PAM is not known.
Mechanism:
1. Ser 226 acts as the primary nucleophile and attacks the amide carbonyl carbon atom.
2. Simultaneously, Ser 202 protonates the substrate carbonyl oxygen, and deprotonates Ser 226. A tetrahedral intermediate is formed.
3. Lys 123 acts to decrease the nucleophilicity of Ser 202, and increase it's ability to protonate the carbonyl oxygen.
4. Lys 123 protonates Ser 202 which in turn protonates the amido group of the substrate, creating NH3, a good leaving group.
5. Lys 123 deprotonates Ser 202 which in turn deprotonates the O atom of the substrate, causing the reformation of the carbonyl with elimination of NH3.
6. Hydrolysis of the enzyme-acyl intermediate is carried out by nucleophilic attack of a water molecule, which is simultaneously deprotonated by the leaving group Ser 226.
Sites:

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Found by:
Literature reference 

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
LYSA 123 123Sidechain
ElectrostaticResidue
Acid/baseResidue
Acts to decrease the nucleophilicity of Ser 202 by hydrogen bonding to it. Also acts as an acid/base to Ser 202.
Evidence from paper Evidence concerns Evidence type
PubMed ID 12367528 Current protein pH dependence of reaction
PubMed ID 12367528 Current protein Kinetic studies
PubMed ID 12367528 Current protein Mutagenesis of residue
PubMed ID 12367528 Current protein Residue is positioned appropriately (ligand position known)

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
SERA 202 202Sidechain
Acid/baseResidue
Acid/baseSubstrate
Acts as an acid/base. Donates a proton to the carbonyl oxygen of the substrate at the same time as abstracting a proton from Ser 226. Accepts a proton from Lys 123 while donating a proton to substrate N atom. Accepts a proton from substrate O atom while donating a proton back to Lys 123.
Evidence from paper Evidence concerns Evidence type
PubMed ID 12367528 Current protein Mutagenesis of residue
PubMed ID 12367528 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 12367528 Current protein Kinetic studies

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
SERA 226 226Sidechain
Acid/baseWater
NucleophileSubstrate
Acts as the primary nucleophile on the substrate carbonyl. Acts as a base by deprotonating a water molecule as it nucleophilically attacks the substrate carbonyl.
Evidence from paper Evidence concerns Evidence type
PubMed ID 12367528 Current protein Mutagenesis of residue
PubMed ID 12367528 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 12367528 Current protein Kinetic studies
Notes:

References:
1
An alternative mechanism for amidase signature enzymes.
J. Labahn and S. Neumann and G. B├╝ldt and M. R. Kula and J. Granzin
J Mol Biol 322, (5) 1053-64, (2002).
12367528
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