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Search The CSA
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Catalytic Site Atlas

CSA LITERATURE entry for 1uaq

E.C. namecytosine deaminase
SpeciesSaccharomyces cerevisiae (Baker's yeast)
E.C. Number (IntEnz) 3.5.4.1
CSA Homologues of 1uaq1ox7,1p6o,1rb7,1teo,1vq2,1ysb,1ysd,2o3k,
CSA Entries With UniProtID Q12178
CSA Entries With EC Number 3.5.4.1
PDBe Entry 1uaq
PDBSum Entry 1uaq
MACiE Entry 1uaq

Literature Report

IntroductionCytosine deaminase catalyses the hydrolytic deamination of cytosine to uracil. It will also catalyse the deamination of the prodrug 5-fluorocytosine. The is present in bacterial and fungal cells, where it plays an important role in pyrimidine salvage, but not in mammalian cells which use cytidine deaminase instead. The bacterial and yeast cytosine deaminases are have dissimilar folds and catalytic site architectures, and have evolved independently. Yeast cytosine deaminase is however structurally related to the extensively studied bacterial cytidine deaminase. It has a very similar catalytic apparatus and is thought to use the same mechanism.
MechansimThe key catalytic residues are Glu 64 and a zinc ion. Glu 64 first deprotonates the zinc-bound water molecule and protonates N3 of cytosine, thus activating both nucleophile and electrophile. Attack by the water molecule on C4 then generates a tetrahedral intermediate. Generation of the tetrahedral intermediate is thought to occur in this stepwise fashion since calculations suggest that a concerted mechanism would have a much higher energy barrier. Collapse of the tetrahedral intermediate involves deprotonation of the zinc-bound C4 hydroxyl followed by cleavage of the C4-N bond with concerted protonation of the departing amino group by Glu 64.
The produced uracil is still coordinated to the zinc by O4. Calculations suggest that the energy barrier for cleavage of this bond is surprisingly high, so it is proposed that freeing of the uracil from the active site involves a gem-diol intermediate and oxygen-exchange. Formation of the gem-diol intermediate involves attack by a water molecule on C4, with concomitant deprotonation of this water molecule by Glu 64. Protonation of the the C4-O-Zn oxygen by Glu 64 is followed by cleavage of the C4-OHZn bond with simulataneous deprotonation of C4-OH by Glu 64.
Reaction

Catalytic Sites for 1uaq

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
GluA6464macie:sideChainDeprotonates the zinc-bound water to give a stronger nucleophile and protonates N3 of cytosine to generate a stronger electrophile. Deprotonates the zinc bound C4 hydroxyl of the intermediate and then protonates the departing C4 amino group. Deprotonates attacking water molecule during formation of the gem-diol intermediate. Protonates the C4-OZn oxygen and then deprotonates the C4-OH during cleavage of the C4-OZn bond.
CysA9191macie:mainChainAmideForms a strong hydrogen bond to the zinc-bound oxygen specifically in the transition state the corresponds to cleavage of the C4-OZn bond in the last stages of the reaction.
SerA8989macie:mainChainCarbonylForms a strong hydrogen bond to the departing amino group specifically in the transition state that corresponds to cleavage of the C4-N bond.

Annotated By Reference To The Literature - Site 2 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
GluB6464macie:sideChainDeprotonates the zinc-bound water to give a stronger nucleophile and protonates N3 of cytosine to generate a stronger electrophile. Deprotonates the zinc bound C4 hydroxyl of the intermediate and then protonates the departing C4 amino group. Deprotonates attacking water molecule during formation of the gem-diol intermediate. Protonates the C4-OZn oxygen and then deprotonates the C4-OH during cleavage of the C4-OZn bond.
CysB9191macie:mainChainAmideForms a strong hydrogen bond to the zinc-bound oxygen specifically in the transition state the corresponds to cleavage of the C4-OZn bond in the last stages of the reaction.
SerB8989macie:mainChainCarbonylForms a strong hydrogen bond to the departing amino group specifically in the transition state that corresponds to cleavage of the C4-N bond.

Literature References

Notes:
Ko TP
Crystal structure of yeast cytosine deaminase. Insights into enzyme mechanism and evolution.
J Biol Chem 2003 278 19111-19117
PubMed: 12637534
Ireton GC
The 1.14 A crystal structure of yeast cytosine deaminase: evolution of nucleotide salvage enzymes and implications for genetic chemotherapy.
Structure 2003 11 961-972
PubMed: 12906827
Carlow DC
Major contribution of a carboxymethyl group to transition-state stabilization by cytidine deaminase: mutation and rescue.
Biochemistry 1995 34 4220-4224
PubMed: 7703234
Sklenak S
Catalytic mechanism of yeast cytosine deaminase: an ONIOM computational study.
J Am Chem Soc 2004 126 14879-14889
PubMed: 15535715
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