A kinetic, modeling and mechanistic re-analysis of thymidine phosphorylase and some related enzymes.
Thymidine phosphorylase (TP) is an important target enzyme for cancer chemotherapy but currently available inhibitors lack in vivo potency. Related enzymes also are therapeutic targets. A greater understanding of enzyme structure and mechanism may help in the design of improved drugs and this work assists in that regard. Also important is the correct identification of the ionization states and tautomeric forms of substrates and products when bound to the enzyme and during the course of the reaction. Approximate methods for estimating some deltapK(a)s between aqueous and protein-bound substrates are exemplified for nucleobases and nucleosides. The estimates demonstrate that carbonyl-protonated thymidine and hydroxy tautomers of thymine are not involved in TP's actions. Other estimates indicate that purine nucleoside phosphorylase binds inosine and guanosine as zwitterionic tautomers and that phosphorolysis proceeds through these forms. Extensive molecular modeling based on an X-ray structure of human TP indicates that TP is likely to be mechanistically similar to all other natural members of the class in proceeding through a alpha-oxacarbenium-like transition state or states.
Edwards
Philip N
PN
School of Pharmacy and Pharmaceutical Sciences, University of Manchester, Oxford Road, Manchester M13 9PL, UK. Philip.N.Edwards@manchester.ac.uk
D000818:Animals; D001665:Binding Sites; D006801:Humans; D007700:Kinetics; D008954:Models, Biological; D010758:Phosphorus / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013939:Thymidine Phosphorylase / Q000737:chemistry / Q000378:metabolism*
D010758:Phosphorus; D013939:Thymidine Phosphorylase