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Catalytic Site Atlas

CSA LITERATURE entry for 1i78

E.C. nameomptin
SpeciesEscherichia coli (Bacteria)
E.C. Number (IntEnz) 3.4.23.49
CSA Homologues of 1i78
CSA Entries With UniProtID P09169
CSA Entries With EC Number 3.4.23.49
PDBe Entry 1i78
PDBSum Entry 1i78
MACiE Entry 1i78

Literature Report

IntroductionOuter membrane protease (OmpT) of Escherichia coli is from a family of homologous outer membrane proteases called omptins. They are implicated in the virulence of several pathogenic, gram negative bacteria, and OmpT is associated with urinary tract disease. OmpT preferentially cleaves substrates between two consecutive basic amino acids, and is dependent on the presence of lipopolysaccharide.
MechansimTheoretical. 1. His 212 activates Ser 99 by acting as a general base. 2. The protonated form of His 212 is stabilised through hydrogen bonding to Asp 210. 3. Ser 99 nucleophilically attacks the scissile peptide bond, forming a negatively charged intermediate. 4. This intermediate is stabilised by the oxyanion hole formed by proton sharing between Asp 83, Asp 85 and possibly a water molecule. 5. When the carbonyl of the substrate is reformed, the leaving group is protonated by His 212. 6. His 212 deprotonates a water molecule which activates it for nucleophilic attack on the carbonyl of the substrate. Another negatively charged, tetrahedral intermediate is formed. 7. On reforming the carbonyl, Ser 99 is the leaving group, and is protonated by His 212.
Reaction

Catalytic Sites for 1i78

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
HisA212232macie:sideChainActivates Ser 99 by acting as a general base. Activates a water molecule again by acting as a general base. Provides leaving group with proton by acting as a general acid.
AspA85105macie:mainChainAmideForms the oxyanion hole and stabilises the negatively charged intermediate via proton sharing (perhaps through a water molecule) with Asp 83.
AspA83103macie:mainChainAmideForms the oxyanion hole and stabilises the negatively charged intermediate via proton sharing (perhaps through a water molecule) with Asp 85.
AlaA99119macie:sideChainHydroxyl side group is deprotonated by His 212. This then goes on to nucleophilically attack the scissile peptide bond of the substrate.
AspA210230macie:sideChainIs thought to either stabilise the protonated form of His 212, or in acting as a base to activate water for nucleophilic attack.

Annotated By Reference To The Literature - Site 2 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
HisB212232macie:sideChainActivates Ser 99 by acting as a general base. Activates a water molecule again by acting as a general base. Provides leaving group with proton by acting as a general acid.
AspB85105macie:mainChainAmideForms the oxyanion hole and stabilises the negatively charged intermediate via proton sharing (perhaps through a water molecule) with Asp 83.
AspB83103macie:mainChainAmideForms the oxyanion hole and stabilises the negatively charged intermediate via proton sharing (perhaps through a water molecule) with Asp 85.
AlaB99119macie:sideChainHydroxyl side group is deprotonated by His 212. This then goes on to nucleophilically attack the scissile peptide bond of the substrate.
AspB210230macie:sideChainIs thought to either stabilise the protonated form of His 212, or in acting as a base to activate water for nucleophilic attack.

Literature References

Notes:This mechanism is no more than a 'best guess'. Ser 99 is proposed to act as a nucleophile, but on mutagenesis, activity is not reduced as much as expected. Also, Ser 99 is not in exactly the place it is expected for a serine protease. The Asp 83 - Asp 85 couple is proposed to be acting as the oxyanion hole, but in fact may be acting as general bases to activate water for nucleophilic attack.
Kramer RA
Identification of active site serine and histidine residues in Escherichia coli outer membrane protease OmpT.
FEBS Lett 2000 468 220-224
PubMed: 10692590
Vandeputte-Rutten L
Crystal structure of the outer membrane protease OmpT from Escherichia coli suggests a novel catalytic site.
EMBO J 2001 20 5033-5039
PubMed: 11566868
Baaden M
OmpT: molecular dynamics simulations of an outer membrane enzyme.
Biophys J 2004 87 2942-2953
PubMed: 15315948
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