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Methods Mol Biol. 2012;848:465-78. doi: 10.1007/978-1-61779-545-9_29.

Flexizymes as a tRNA acylation tool facilitating genetic code reprogramming.

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1
Department of Chemistry, Graduate School of Science, The University of Tokyo, Tokyo, Japan.

Abstract

Genetic code reprogramming is a method for the reassignment of arbitrary codons from proteinogenic amino acids to non-proteinogenic ones, and thus specific sequences of nonstandard peptides can be ribosomally expressed according to their mRNA templates. We here describe a protocol that facilitates the genetic code reprogramming using flexizymes integrated with a custom-made in vitro translation apparatus, referred to as the flexible in vitro translation (FIT) system. Flexizymes are flexible tRNA acylation ribozymes that enable the preparation of a diverse array of non-proteinogenic acyl-tRNAs. These acyl-tRNAs read vacant codons created in the FIT system, yielding the desired nonstandard peptides with diverse exotic structures, such as N-acyl groups, D: -amino acids, N-methyl amino acids, and physiologically stable macrocyclic scaffolds. Facility of the protocol allows for a wide variety of applications in the synthesis of new classes of nonstandard peptides with biological functions.

PMID:
22315087
DOI:
10.1007/978-1-61779-545-9_29
[Indexed for MEDLINE]
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