Nucleic Acids Research, 2000, Vol. 28, No. 23 4725-4732
© 2000 Oxford University Press
Sequestration of specific tRNA species cognate to the last sense codon of an overproduced gratuitous protein
UPR9073 du CNRS, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, F-75005 Paris, France
High-level expression of non-functional model proteins, derived from elongation factor EF-Tu by the deletion of an essential domain, greatly inhibits the growth of Escherichia coli partly deficient in peptidyl-tRNA hydrolase. High-level expression in wild-type cells has little effect on growth. The inhibitory effect is therefore presumably due to the sequestration of essential tRNA species, partly in the form of free peptidyl-tRNA. The growth inhibitory effect can be modulated by changing the last sense codon in the genes encoding the model proteins. Thus, replacement of Ser by Lys or His at this position increases growth inhibition. The effects of 11 changes studied are related to the rates of accumulation previously observed of the corresponding families of peptidyl-tRNA. Two non-exclusive hypotheses are proposed to account for these observations: first, the last sense codon of mRNA is a prefered site of peptidyl-tRNA drop-off in cells, due to the slow rate of translation termination compared with sense codon translation; secondly, the relatively long pause of the ribosome at the stop codon (of the order of 1 s), results in significant temporary sequestration on the ribosome of the tRNA cognate to the last sense codon.
* To whom correspondence should be addressed. Tel: +33 1 4354 1972; Fax: +33 1 4046 8331; Email: rhb{at}ibpc.fr The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors
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