Kinetic characterization of bifunctional thymidylate synthase-dihydrofolate reductase (TS-DHFR) from Cryptosporidium hominis: a paradigm shift for ts activity and channeling behavior |
| |
Authors: | Atreya Chloé E Anderson Karen S |
| |
Institution: | Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520, USA. |
| |
Abstract: | This study presents a kinetic characterization of the recently crystallized bifunctional thymidylate synthasedihydrofolate reductase (TS-DHFR) enzyme from the apicomplexa parasite, Cryptosporidium hominis. Our study focuses on determination of the C. hominis TS-DHFR kinetic mechanism, substrate channeling behavior, and domain-domain communication. Unexpectedly, the unique mechanistic features of C. hominis TS-DHFR involve the highly conserved TS domain. At 45 s(-) (1), C. hominis TS activity is 10-40-fold faster than other TS enzymes studied and a new kinetic mechanism was required to simulate C. hominis TS behavior. A large accumulation of dihydrofolate produced at TS and a lag in product formation at DHFR were observed. These observations make C. hominis TS-DHFR the first bifunctional TS-DHFR enzyme studied for which there is clear evidence against dihydrofolate substrate channeling. Furthermore, whereas with Leishmania major TS-DHFR there are multiple lines of evidence for domain-domain communication (ligand binding at one active site affecting activity of the other enzyme), no such effects were observed with C. hominis TS-DHFR. |
| |
Keywords: | |
本文献已被 PubMed 等数据库收录! |
|