High-Throughput Mutational Analysis of a Methyltransferase Ribozyme

Methyltransferase ribozyme 1 (MTR1) is a catalytic RNA that was discovered from a random RNA pool by in vitro selection. The ribozyme catalyzes a site-specific 1-methyl adenosine (m1A) formation using 6-methyl guanine (m6G) base as a methyl group donor. Characteristic properties of the ribozyme have been extensively characterized by biochemical and structural analyses. While broad applications of the methyltransferase ribozyme have been expected, a high-throughput screening method for the measurement of the ribozyme activity has not been established yet. In this study, we apply the mutational profiling technique to the methyltransferase assay where we analyze the activities in 141 distinct MTR1 variants in which single-point variations are introduced in the catalytic core domain. Our method successfully detected the activity in the all MTR1 variants tested in a high-throughput manner.