The isolation of methyl deficient phenylalanyl transfer RNA from [formula omitted][formula omitted

Abstract

The study of the role of methylated bases in transfer RNA (t-RNA) has been made possible by the discovery by Mandel and Borek (1961) that methyl deficient s-RNA is synthesized by a “relaxed” methionine requiring mutant of E.coli during methionine starvation. Preparations of s-RNA from methionine starved cells and from normal cells have been compared but no significant differences in the ability of these RNA's to accept (Starr, 1963; Littauer et al., 1963; Peterkofsky et al. 1964) and to transfer amino acids (Littauer et al., 1963; Peterkofsky et al. 1964) have been detected. Although this may indicate that the methylated bases do not participate in these functions of t-RNA, it should be recalled that “methionine starved” RNA preparations consist of a mixture of equal amounts of methyl deficient and normal s-RNA. Therefore, small differences in the activities of these RNA species invitro, which could be very critical invivo, might not have been detected with such crude RNA preparations. These considerations prompted us to develop a method for the isolation of methyl deficient t-RNA. Phenylalanyl transfer RNA (phe-t-RNA) from E.coli G-15, a relaxed double mutant which can be starved for methionine or for histidine, was selected for these studies. The present report describes the isolation of methyl deficient phe-t-RNA by chromatography on methylated albumin kieselguhr (MAK) columns.