Enzymatic synthesis of nucleoside antibiotics. Part VI. Enzymatic synthesis of virazole by purine nucleoside phosphorylase of Enterobacter aerogenes.

Abstract

A potent antiviral agent, Virazole (1-β-D-ribofuranosyl-1, 2, 4-triazole-3-carboxamide), was synthesized from inosine and 1, 2, 4-triazole-3-carboxamide (TCA) by a two step reaction with purified purine nucleoside phosphoryiase (PNPase) of Enterobacter aerogenes AJ 11125. The first step involves the production of ribose-1-phosphate (R-1-P) through the phosphorolysis of inosine. The second step involves the production of Virazole through the transribosyl reaction of R-1-P and TCA. The R-1-P produced from inosine was isolated by ion-exchange chromatography and used as a substrate for Virazole synthesis without crystallization. TCA (10 mM) was transformed to Virazole with a 75% yield (molar basis) in the presence of 50mM R-1-P. Enzymatically synthesized Virazole was isolated from a large scale reaction mixture and identified by physicochemical means. The Km values of PNPase for TCA and hypoxanthine were 167 mM and 5.6 mM, respectively.