Journal
FEBS LETTERS
Volume 579, Issue 10, Pages 2156-2160Publisher
WILEY
DOI: 10.1016/j.febslet.2005.03.004
Keywords
in vitro translation; suppression; tRNA; release factor 1; esterase; termination
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An mRNA encoding the esterase from Alicyclobacillus acidocaldarius with catalytically essential serine codon (ACG) replaced by an amber (UAG) codon was used to study the suppression in in vitro translation system. Suppression of UAG by tRNA (Ser(CUA)) was monitored by determination of the full-length and active esterase. It was shown that commonly used increase of suppressor tRNA concentration inhibits protein production and therefore limits suppression. In situ deactivation of release factor by specific antibodies leads to efficient suppression already at low suppressor tRNA concentration and allows an in vitro synthesis of fully active enzyme in high yield undistinguishable from wild-type protein. (c) 2005 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
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