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Catalytic Site Atlas

CSA LITERATURE entry for 1x9y

E.C. nameCYSTEINE PROTEINASE
SpeciesStaphylococcus aureus (Bacteria)
E.C. Number (IntEnz) 3.4.22.-
CSA Homologues of 1x9y
CSA Entries With UniProtID
CSA Entries With EC Number 3.4.22.-
PDBe Entry 1x9y
PDBSum Entry 1x9y
MACiE Entry 1x9y

Literature Report

IntroductionStaphopain B (SspB)is a papain-type secreted cysteine protease from the pathogen Staphylococcus aureus. It is a produced as an inactive proenzyme, which is comprised of the active part of the enzyme, the mature form, and a covalently bonded inhibitory proregion. Activation of the enzyme requires cleavage of the N terminal inhibitory proregion (staphostatin B).
MechansimThe overall mechanism involves a nucleophilic cysteine thiol in a catalytic triad.
1. Deprotonation of the Cys 243 sulfhydryl by an adjacent histidine residue with a basic side chain. The thiolate ion is stabilised through the formation of an ion pair with the neighbouring, approximately coplanar imidazolium group of His 340. The carbonyl bond of the scissile peptide bond in the substrate is directed towards an oxyanion hole formed from the main chain Cys NH and a Gln 237 amide side chain. The ion pair is stabilised by Gln 237 and Asn 360. Asn 360 is adjacent to the catalytic His 340, and its side chain amide oxygen is hydrogen bonded to the N(e2)H of His 340. This effect of this is to both stabilise the ion pair and also keep the imidazole ring of the His residue in favourable orientation.
2. Nucleophilic attack of the anionic cysteine S (thiolate ion) on the peptide carbonyl carbon. In this step, a fragment of the substrate is released with an amine terminus, the histidine residue in the protease is restored to its deprotonated form, and a thioester intermediate linking the new carboxy-terminus of the substrate to the cysteine thiol is formed.
3. The thioester bond is subsequently hydrolysed to generate a carboxylic acid moiety on the remaining substrate fragment, whilst regenerating the free enzyme.

Catalytic Sites for 1x9y

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
HisA340340macie:sideChainThe basic side chain of the His residue deprotonates the cysteine thiol to activate it towards nucleophilic attack of the substrate peptide bond. The imidazole ring of His 340 forms an ion pair with Cys 243. His 340 forms a hydrogen bond to the Asn 360 side chain amide oxygen to stabilise the ion pair and keep the imidazole ring in favourable orientation.
CysA243243macie:sideChainDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
CysA243243macie:mainChainAmideDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
GlnA237237macie:sideChainThe Gln 237 amide side chain forms an oxyanion hole, along with the Cys 243 main chain NH. This stabilises the transition state.
AsnA360360macie:sideChainAsn 360 is adjacent to the catalytic His 340, and its side chain amide oxygen is hydrogen bonded to the N(e2)H of His 340. This effect of this is to both stabilise the ion pair and also keep the imidazole ring of the His residue in favourable orientation.

Annotated By Reference To The Literature - Site 2 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
HisB340340macie:sideChainThe basic side chain of the His residue deprotonates the cysteine thiol to activate it towards nucleophilic attack of the substrate peptide bond. The imidazole ring of His 340 forms an ion pair with Cys 243. His 340 forms a hydrogen bond to the Asn 360 side chain amide oxygen to stabilise the ion pair and keep the imidazole ring in favourable orientation.
CysB243243macie:sideChainDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
CysB243243macie:mainChainAmideDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
GlnB237237macie:sideChainThe Gln 237 amide side chain forms an oxyanion hole, along with the Cys 243 main chain NH. This stabilises the transition state.
AsnB360360macie:sideChainAsn 360 is adjacent to the catalytic His 340, and its side chain amide oxygen is hydrogen bonded to the N(e2)H of His 340. This effect of this is to both stabilise the ion pair and also keep the imidazole ring of the His residue in favourable orientation.

Annotated By Reference To The Literature - Site 3 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
HisC340340macie:sideChainThe basic side chain of the His residue deprotonates the cysteine thiol to activate it towards nucleophilic attack of the substrate peptide bond. The imidazole ring of His 340 forms an ion pair with Cys 243. His 340 forms a hydrogen bond to the Asn 360 side chain amide oxygen to stabilise the ion pair and keep the imidazole ring in favourable orientation.
CysC243243macie:sideChainDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
CysC243243macie:mainChainAmideDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
GlnC237237macie:sideChainThe Gln 237 amide side chain forms an oxyanion hole, along with the Cys 243 main chain NH. This stabilises the transition state.
AsnC360360macie:sideChainAsn 360 is adjacent to the catalytic His 340, and its side chain amide oxygen is hydrogen bonded to the N(e2)H of His 340. This effect of this is to both stabilise the ion pair and also keep the imidazole ring of the His residue in favourable orientation.

Annotated By Reference To The Literature - Site 4 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
HisD340340macie:sideChainThe basic side chain of the His residue deprotonates the cysteine thiol to activate it towards nucleophilic attack of the substrate peptide bond. The imidazole ring of His 340 forms an ion pair with Cys 243. His 340 forms a hydrogen bond to the Asn 360 side chain amide oxygen to stabilise the ion pair and keep the imidazole ring in favourable orientation.
CysD243243macie:sideChainDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
CysD243243macie:mainChainAmideDeprotonation of the cysteine thiol by the His 340 basic side chain activates the cysteine S to carry out nucleophilic attack on the carbonyl carbon of the peptide bond in the substrate. The thiolate ion is stabilised by the formation of an active site ion pair with the His 340 imidazole ring. The main chain NH of Cys 243 forms part of the oxyanion hole, which stabilises the transition state.
GlnD237237macie:sideChainThe Gln 237 amide side chain forms an oxyanion hole, along with the Cys 243 main chain NH. This stabilises the transition state.
AsnD360360macie:sideChainAsn 360 is adjacent to the catalytic His 340, and its side chain amide oxygen is hydrogen bonded to the N(e2)H of His 340. This effect of this is to both stabilise the ion pair and also keep the imidazole ring of the His residue in favourable orientation.

Literature References

Notes:The reaction has found to be concerted, rather than stepwise, via the formation of a tetrahedral transition state.
M. J. Harrison
Catalytic Mechanism of the Enzyme Papain: Predictions with a Hybrid Quantum Mechanical/Molecular Mechanical Potential
J. Am. Chem. Soc. 1997 119 12285-12291
PubMed: Harrison1997
Filipek R
The Staphostatin-staphopain complex: a forward binding inhibitor in complex with its target cysteine protease.
J Biol Chem 2003 278 40959-40966
PubMed: 12874290
Filipek R
Prostaphopain B structure: a comparison of proregion-mediated and staphostatin-mediated protease inhibition.
Biochemistry 2004 43 14306-14315
PubMed: 15518582
Filipek R
A comparison of staphostatin B with standard mechanism serine protease inhibitors.
J Biol Chem 2005 280 14669-14674
PubMed: 15644332
Ballistreri A
Biosynthesis and structural characterization of medium-chain-length poly(3-hydroxyalkanoates) produced by Pseudomonas aeruginosa from fatty acids.
Int J Biol Macromol 2001 29 107-114
PubMed: 11518582
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