Purification of wheat germ guanine insertase and observations using an X-irradiated transfer RNA substrate

Guanine insertase has been purified 3382-fold to homogeneity with a specific activity of 2,029 pmoles hr -1 The enzyme, which has a molecular weight of l40,000, migrates as a single band on polyacrylamide gel electrophoresis. The enzyme substitutes guanine for guanine and is inhibited by Q base. A monovalent cation best met by are required for guanylation. . The divalent metal obliviates the monovalent metal requirement. The K for guanine is 1.05 x 10-6M and the optimum pH is 7.6. Seven tRNAs found in wheat germ are substrates for guanine insertase.

Observations on the guanine accepting ability of irradiated wheat germ tRNA shows 50% inactivation at 60,000 Roentgens with an unusual saw tooth pattern at low doses. The saw tooth has peaks at 2,000 and 4,00o Roentgens and can be modified by Tris or bovine serum albumin. RPC5 profiles of guanylated irradiated and unirradiated tRNA are identical.