STUDY ON CATALYTIC REACTION MECHANISM OF ADENOSYLHOMOCYSTEINE HYDRLASE AND SYNTHESIS OF THEIR MECANISM -BASED INHIBITORS

The cellular enzyme S-adenosyl-L-homocysteine(SAH)hydrolase(EC3.3.1.1)has emerged as a target enzyme for the molecular design of anti-viral agents.During the course of studies on the preparation of biologically important nucleosides using the reductive cleavage of purine nucleosides，a DHPA analogue(FDHPA)possessing a formyl group at the 3‘-position has been designed as a possible affinity-labeling probe for the elucidation of the catalytic site of SAH hydrolase。We describe a method for the preparation of a facile affinity-labeling probe and its biological properties against human recombinant SAH hydrolase.The cDNA for human SAH hydrolase has been cloned from human heparoma cell HepG2 by RT-PCR and the obtained human recombinant SAH hydrolase was purified according to the literature.Incubation with FDHPA caused inactivation of the human SAH hydrolase and the activity was not recovered by dialysis The values of K i-D2i-D2 and k-D2inact D2were 8.8µM and 0.09min I D1-1 I D1，respectively。Taking the above results into consideration，it is deduced that FDHPA functions as a useful affinity-labeling probe of SAH hydrolase and provides a clue to elucidation of the molecular mechanism of SAH hydrolase aiming at the molecular design of anti-viral drugs.We also prepared several carbocyclic nucleosides and acyclic nucleosides for the discovery of now type SAH drolinhibitors.

The cellular enzyme S-adenosyl-L-homocysteine (SAH) hydrolase (EC 3.3.1.1) has emerged as a target enzyme for the molecular design of anti-viral agents. During the course of studies on the preparation of biologically important nucleosides using the reductive cleavage of purine nucleosides, a DHPA analogue (FDHPA) possessing a formyl group at the 3'-position has been designed as a possible affinity-labeling probe for the elucidation of the catalytic site of SAH hydrolase. We describe a method for the preparation of a facile affinity-labeling probe and its biological properties against human recombinant SAH hydrolase.The cDNA for human SAH hydrolase has been cloned from human heparoma cell HepG2 by RT-PCR and the obtained human recombinant SAH hydrolase was purified according to the literature. Incubation with FDHPA caused inactivation of the human SAH hydrolase and the activity was not recovered by dialysis The values of KィイD2iィエD2 and kィイD2inactィエD2 were 8.8 μM and 0.09 minィイD1-1ィエD1, respectively. Taking the above results into consideration, it is deduced that FDHPA functions as a useful affinity-labeling probe of SAH hydrolase and provides a clue to elucidation of the molecular mechanism of SAH hydrolase aiming at the molecular design of anti-viral drugs.We also prepared several carbocyclic nucleosides and acyclic nucleosides for the discovery of now type SAH hydrolase inhibitors.