Deoxynucleotide-polymerizing Enzymes of Calf Thymus Gland

作者：Kenichi Kato, J. Moura Gonalves, G. E. Houts, F.J. Bollum · 发表于：Journal of Biological Chemistry · 年份：1967 · DOI：10.1016/s0021-9258(18)99635-3 · 被引用次数：324 · 研究领域：biodegradable polymer synthesis and properties、Biochemical and Molecular Research、HIV/AIDS drug development and treatment

Abstract The stoichiometry, equilibrium, and kinetic properties of a terminal deoxynucleotidyltransferase isolated from calf thymus gland are described. The reaction catalyzed may be written dXTP + d(pY)m (Mg++)/→ d(pY)m(pX)n + nPPi and the ratio of polymer nucleotide to monomer at equilibrium (pH 7.0, 35°) is about 99 when X and Y = adenine. Purine deoxyribonucleoside triphosphate polymerizations proceed readily in 8 mm Mg++ at low buffer strengths (40 mm). For deoxyadenosine triphosphate, under these conditions, Km = 1.1 x 10-4 m and Vmax varies from about 0.03 to 0.30 µmole min-1 mg-1 with various oligodeoxynucleotide initiators. At a constant chain length, Km values for oligodeoxynucleotide decrease in the order cytosine g thymine g adenine g guanine g hypoxanthine. Within a homologous series, a limiting value of Km and a maximum value for Vmax are reached at the penta- to septanucleotide. Pyrimidine deoxyribonucleoside triphosphates polymerize best in the presence of 1 mm Co++ at higher buffer strengths (200 mm). Syntheses of polydeoxyadenylate, polydeoxyinosinate, polydeoxycytidylate, and polydeoxythymidylate in greater than 80% yield are achieved under optimal conditions. Preparation of random copolymers is also described.