Deoxyribozyme-based, semisynthetic access to stable peptidyl-tRNAs exemplified by tRNA Val carrying a macrolide antibiotic resistance peptide

Abstract

We present a protocol for the reliable synthesis of non-hydrolyzable 3?-peptidyl-tRNAs that contain all the respective genuine nucleoside modifications. The approach is exemplified by tRNA Val-3?-NH-VFLVM- NH 2 and relies on commercially available Escherichia coli tRNA Val. This tRNA was cleaved site-specifically within the TΨC loop using a 10-23 type DNA enzyme to obtain a 58 nt tRNA 5?-fragment which contained the modifications. After cleavage of the 2?,3?-cyclophosphate moiety from the 5?-fragment, it was ligated to the 18 nt RNA-pentapeptide conjugate which had been chemically synthesized. By this methodology, tRNA Val-3′- NH-VFLVM-NH 2 is accessible in efficient manner. Furthermore, we point out that the approach is applicable to other types of tRNA. © 2012 Springer Science+Business Media, LLC.