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Puromycin reaction for the A site‐bound peptidyl‐tRNA
Author(s) -
Semenkov Yu.,
Shapkina T.,
Makhno V.,
Kirillov S.
Publication year - 1992
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(92)80380-y
Subject(s) - puromycin , chemistry , transfer rna , stereochemistry , biochemistry , protein biosynthesis , rna , gene
AcPhe 2 ‐tRNA Phe synthesized in 70S ribosomes after consecutive binding of AcPhe‐tRNA Phe at the P sites and EF‐Tu‐directed binding of Phe‐tRNA Phe at the A sites is able to react quantitatively with puromycin in the absence of EF‐G. A detailed study of the kinetics of the puromycin reaction, its comparison with that of spontaneous translocation, the use of antibiotic viomycin as an effective inhibitor of spontaneous translocation revealed that, besides spontaneous translocation, this peptidyl‐tRNA could react with puromycin being located at the A site. This leads to the conclusion that the transpeptidation reaction per se triggers conformational changes in the ribosomal complex bringing the 3′‐end of a newly synthesized peptidyl‐tRNA nearer to the peptidyl‐site of the peptidyltransferase center. This is detected functionally as the ability of such an A site bound peptidyl‐tRNA to react with puromycin. This reaction is highly pronounced at elevated (25°C) temperature but can be hardly detected at 0°C.