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A cryptic ribosome binding site, false signals in reporter systems and avoidance of protein translation chaos

Paper ID Volume ID Publish Year Pages File Format Full-Text
24498 43520 2009 4 PDF Available
Title
A cryptic ribosome binding site, false signals in reporter systems and avoidance of protein translation chaos
Abstract

The expression of reporter gene may be induced by activation of cryptic signalling sequences, as we found while constructing the mutS-lacZ fusion gene. We cloned the Escherichia coli lacZ gene encoding β-galactosidase into a plasmid vector carrying the Thermus thermophilus mutS gene. The clones expected to produce β-galactosidase as the C-terminal fusion were selected for the complementation of β-galactosidase activity in a lacZ deficient E. coli strain. Surprisingly, one of the clones, though displaying β-galactosidase activity, did not produce the fusion protein. As shown by DNA sequencing a 92 bp fragment in the 3′ part of mutS gene was substituted by a 19 bp sequence. As the consequence of the resultant frameshift, a truncated MutS peptide was translated instead of β-galactosidase fusion. The cloned lacZ gene lacked its ribosome binding site, so lacZ expression could be explained by activation of a cryptic ribosome binding site in the 3′ end of mutS gene. This observation shows that fusion domains in reporter systems are possible to produce accidentally misleading signals. This observation also suggests that some triplets like AGG and AGA, present in the canonical ribosome binding sequence, are rarely used codons to prevent chaotic protein translation.

Keywords
IPTG, isopropyl-beta-d-thiogalactopyranoside; rbs, ribosome binding site; X-Gal, 5-bromo-4-chloro-3-indolyl-beta-d-galactopyranosideMutS; β-Galactosidase; Fusion; Chimeric protein; Reporter system; Ribosome binding sequence; Rare codons; Translation initi
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A cryptic ribosome binding site, false signals in reporter systems and avoidance of protein translation chaos
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 143, Issue 3, 10 September 2009, Pages 169–172
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us