Biosynthesis of 1-methyladenine by isolated segments of starfish ovaries

Acid-soluble 1-methyladenine (1-MeA) and an insoluble fraction containing 1-MeA were formed when radioactive L-methionine or adenine (A) were incubated with starfish ovary segments. Attempts to prepare free ribonucleic acid (RNA) from ovaries failed since it was strongly bonded to protein as ribonucleoprotein (RN-P) which was therefore used in studies involving RNA. Incubation of ovary segments with [8-14C]adenine-8 (A-8-14C) yielded both soluble 1-Me-A-8-14C and RN-P-8-14C, and similar incubation with L-[methyl-14C]methionine yielded soluble 1-MeA-14CH3 and RN-P-14CH3. Hydrolysis of RN-P-8-14C with 1 N HCl at 100 degrees yielded 91% of the initial radioactivity in the purine fraction, and of this 90% was in A, 9% in 1-MeA, and 1% in 4(5)-amino-5(4)-imidazole carboxamidine (AIMCAD). With RN-P-14CH3 the corresponding figures were 20, 45, 27, and 28%. Degradation of 1-MeA-8-14C or RN-P-8-14C with 6 N HCl at 110 degrees yielded radioactive AIMCAD which, on heating at pH 12, gave radioactive 4(5)-amino-5(4)-imidazole carboxamide (AICA). When 1-MeA-14CH3 or RN-P-14CH3 were similarly degraded, radioactive AIMCAD was formed, but the AICA possessed little or no radioactivity due to the loss of the radioactive methyl group. Addition of radial nerve factor (GSS) increased the yield of 1-MeA up to 19 times when radioactive L-methionine was substrate, but was ineffective with radioactive A. S-[8-14C]Adenosylmethionine was only about 3% as effective as L-[methyl-14C]methionine in supporting formation of 1-MeA, and its slight activity was not enhanced by GSS. Immature ovary segments were much more active than those from mature ovaries in synthesizing 1-MeA. The results support the contention that formation of free 1-MeA in starfish ovarian tissues involves methylation of adenine residues in a polynucleotide followed by liberation of 1-MeA by enzymatic hydrolysis.