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Catalytic Site Atlas

CSA LITERATURE entry for 1cs1

E.C. namecystathionine gamma-synthase
SpeciesEscherichia coli (Bacteria)
E.C. Number (IntEnz) 2.5.1.48
CSA Homologues of 1cs1There are 429 Homologs
CSA Entries With UniProtID P00935
CSA Entries With EC Number 2.5.1.48
PDBe Entry 1cs1
PDBSum Entry 1cs1
MACiE Entry 1cs1

Literature Report

IntroductionCystathionine gamma-synthase (CGS) is a pyrodxial phosphate-dependent enzyme that catalyses a gamma-replacement reaction, in which the succinyl group of an O-succinyl-L-homoserine (L-OSHS) is displaced by the thiol of L-cysteine to form L-cystathionine, in the first step of the bacterial transsulphuration pathway.
CGS is of interest as a potential target for antibiotics and herbicides.
MechansimThe reaction proceeds via a series of steps, as is thought to follow a ping-pong mechanism, commonly encountered in PLP-dependent enzymes.
1. Transaldimination i) OSHS binds through Arg 48*, Tyr 101 and Arg 361. ii) The alpha-amino group of the substrate must be deprotonated for nucleophilic attack on C4' of the internal aldimine. Tyr 101 exists as phenolate due to two neighbouring positive charges (Arg 48* and NH of the internal aldimine). Therefore, Tyr 101 abstracts a proton from the incoming substrate and initiates transaldimination.
2. Generation of the ketimine intermediate i) Lys 198 is responsible for proton transfer from the alpha-C to C4' of the PLP cofactor. The protonated amino group of Lys 198 is guided into a favourable position near C4' by Tyr 46*. After alpha-C deprotonation, a quinonoid intermediate is formed, which is stabilised by stacking interactions with Tyr 101. ii) The Lys 198 e-amino group is positively charged and is therefore attracted to the negatively charged phosphate group of the PLP cofactor, orientating it into a favourable position for bond cleavage. iii) Due to the new positioning of Lys 198, this residue is able to abstract a proton from the beta-C to initiate gamma-cleavage.
3. Release of succinate Tyr 101 facilitates the release of succinate by and acid/base mechanism.
4. The resulting beta-gamma unsaturated ketimine exhibits pronounced electron deficiency, caused by the protonated Schiff base, leading to activation of gamma-C towards Michael nucleophilic addition by L-cysteinate.
5. The reverse steps of 1-3 occur (beta-C protonation, C4' deprotonation, alpha-C protonation.)
Reaction

Catalytic Sites for 1cs1

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
LlpA198198macie:ptmLys 198 acts as a Schiff base with a protonated N, and is stabilised by interactions with the deprotonated PLP cofactor group at C3. Lys 198 abstracts a proton from the beta-C to initiate gamma-cleavage.
TyrA101101macie:sideChainThe phenol group of Tyr 101 exists as phenolate, due to two neighbouring positive charges (Arg 48* and NH of the internal aldimine). Tyr 101 impedes vertical movement of PLP with respect to the pyridine ring since the phenol Tyr ring is positioned appropriately above the PLP pyridine to restrict the cofactor. The resulting stacking interactions also increase the electron sink character of the PLP cofactor. Tyr 101 abstracts a proton from the incoming substrate to initiate transaldimination and also faciliates the release of succinate by and acid/base mechanism.
AspA173173macie:sideChainAsp 173 forms a strong hydrogen bond to the PLP pyridine N1, therefore stabilising its positive charge at this position, and increasing the electrophilic character of the cofactor. The carboxylate group of Asp 173 is fixed in the geometrically optimal position for contact with the substrate N1.
ArgC4848macie:sideChainArg 48* allows binding of the PLP phophate and interaction with Tyr 101. Therefore increases the nucleophilicity of the Tyr residue.

Annotated By Reference To The Literature - Site 2 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
ArgA4848macie:sideChainArg 48* allows binding of the PLP phophate and interaction with Tyr 101. Therefore increases the nucleophilicity of the Tyr residue.
AspC173173macie:sideChainAsp 173 forms a strong hydrogen bond to the PLP pyridine N1, therefore stabilising its positive charge at this position, and increasing the electrophilic character of the cofactor. The carboxylate group of Asp 173 is fixed in the geometrically optimal position for contact with the substrate N1.
TyrC101101macie:sideChainThe phenol group of Tyr 101 exists as phenolate, due to two neighbouring positive charges (Arg 48* and NH of the internal aldimine). Tyr 101 impedes vertical movement of PLP with respect to the pyridine ring since the phenol Tyr ring is positioned appropriately above the PLP pyridine to restrict the cofactor. The resulting stacking interactions also increase the electron sink character of the PLP cofactor. Tyr 101 abstracts a proton from the incoming substrate to initiate transaldimination and also faciliates the release of succinate by and acid/base mechanism.

Annotated By Reference To The Literature - Site 3 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
AspD173173macie:sideChainAsp 173 forms a strong hydrogen bond to the PLP pyridine N1, therefore stabilising its positive charge at this position, and increasing the electrophilic character of the cofactor. The carboxylate group of Asp 173 is fixed in the geometrically optimal position for contact with the substrate N1.
TyrD101101macie:sideChainThe phenol group of Tyr 101 exists as phenolate, due to two neighbouring positive charges (Arg 48* and NH of the internal aldimine). Tyr 101 impedes vertical movement of PLP with respect to the pyridine ring since the phenol Tyr ring is positioned appropriately above the PLP pyridine to restrict the cofactor. The resulting stacking interactions also increase the electron sink character of the PLP cofactor. Tyr 101 abstracts a proton from the incoming substrate to initiate transaldimination and also faciliates the release of succinate by and acid/base mechanism.
ArgB4848macie:sideChainArg 48* allows binding of the PLP phophate and interaction with Tyr 101. Therefore increases the nucleophilicity of the Tyr residue.

Annotated By Reference To The Literature - Site 4 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
ArgD4848macie:sideChainArg 48* allows binding of the PLP phophate and interaction with Tyr 101. Therefore increases the nucleophilicity of the Tyr residue.
TyrB101101macie:sideChainThe phenol group of Tyr 101 exists as phenolate, due to two neighbouring positive charges (Arg 48* and NH of the internal aldimine). Tyr 101 impedes vertical movement of PLP with respect to the pyridine ring since the phenol Tyr ring is positioned appropriately above the PLP pyridine to restrict the cofactor. The resulting stacking interactions also increase the electron sink character of the PLP cofactor. Tyr 101 abstracts a proton from the incoming substrate to initiate transaldimination and also faciliates the release of succinate by and acid/base mechanism.
AspB173173macie:sideChainAsp 173 forms a strong hydrogen bond to the PLP pyridine N1, therefore stabilising its positive charge at this position, and increasing the electrophilic character of the cofactor. The carboxylate group of Asp 173 is fixed in the geometrically optimal position for contact with the substrate N1.

Literature References

Notes:
Clausen T
Crystal structure of Escherichia coli cystathionine gamma-synthase at 1.5 A resolution.
EMBO J 1998 17 6827-6838
PubMed: 9843488
Clausen T
Crystal structure of the pyridoxal-5'-phosphate dependent cystathionine beta-lyase from Escherichia coli at 1.83 A.
J Mol Biol 1996 262 202-224
PubMed: 8831789
Aitken SM
Escherichia coli cystathionine gamma-synthase does not obey ping-pong kinetics. Novel continuous assays for the elimination and substitution reactions.
Biochemistry 2003 42 11297-11306
PubMed: 14503880
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