Using a solid-phase ribozyme aminoacylation system to reprogram the genetic code.
Here, we report a simple and economical tRNA aminoacylation system based upon a resin-immobilized ribozyme, referred to as Flexiresin. This catalytic system features a broad spectrum of activities toward various phenylalanine (Phe) analogs and suppressor tRNAs. Most importantly, it allows users to perform the tRNA aminoacylation reaction and isolate the aminoacylated tRNAs in a few hours. We coupled the Flexiresin system with a high-performance cell-free translation system and demonstrated protein mutagenesis with seven different Phe analogs in parallel. Thus, the technology developed herein provides a new tool that significantly simplifies the procedures for the synthesis of aminoacyl-tRNAs charged with nonnatural amino acids, which makes the nonnatural amino acid mutagenesis of proteins more user accessible.
Murakami
Hiroshi
H
Department of Chemistry, University at Buffalo, The State University of New York, Buffalo, NY 14260, USA.
Kourouklis
Dimitrios
D
Suga
Hiroaki
H
D000215:Acylation; D001483:Base Sequence; D002384:Catalysis; D002474:Cell-Free System; D017931:DNA Primers; D005815:Genetic Code; D016297:Mutagenesis, Site-Directed; D011499:Protein Processing, Post-Translational; D016337:RNA, Catalytic / Q000235:genetics / Q000378:metabolism*; D012348:RNA, Transfer, Asn / Q000235:genetics / Q000378:metabolism
D017931:DNA Primers; D016337:RNA, Catalytic; D012348:RNA, Transfer, Asn