Purification of post-transcriptionally modified tRNAs for enhanced cell-free translation systems.

ABSTRACT: Transfer RNAs (tRNAs) are utilized by the ribosome to decode the nucleic acid alphabet. tRNA structure, stability, aminoacylation efficiency, and decoding efficacy are governed by their extensive post-transcriptional modifications. In most studies, individual tRNAs are generated using in vitro transcription, which produces tRNAs devoid of these critical site-specific modifications, negatively affecting translation yields and fidelity. To address this, we have developed a purification method which couples tRNA overexpression to DNA hybridization-based purification. Using this approach, we produced native tRNAs from E. coli in high yield and purity while retaining their complement of native post-transcriptional modifications and translational activity. We extend this technique

SUBMITTER: Kalb EM 

PROVIDER: S-EPMC12259135 | biostudies-literature | 2025 Jun

REPOSITORIES: biostudies-literature