The Effects of Codon Context on In Vivo Translation Speed

Abstract

We developed a bacterial genetic system based on translation of the his operon leader peptide gene to determine the relative speed at which the ribosome reads single or multiple codons in vivo. Low frequency effects of so-called “silent” codon changes and codon neighbor (context) effects could be measured using this assay. An advantage of this system is that translation speed is unaffected by the primary sequence of the His leader peptide. We show that the apparent speed at which ribosomes translate synonymous codons can vary substantially even for synonymous codons read by the same tRNA species. Assaying translation through codon pairs for the 5′- and 3′- side positioning of the 64 codons relative to a specific codon revealed that the codon-pair orientation significantly affected in vivo translation speed. Codon pairs with rare arginine codons and successive proline codons were among the slowest codon pairs translated in vivo. This system allowed us to determine the effects of different factors on in vivo translation speed including Shine-Dalgarno sequence, rate of dipeptide bond formation, codon context, and charged tRNA levels.

Synthetic gene design—The rationale for codon optimization and implications for molecular pharming in plants

“…The results indicated that codon context is a major factor affecting translational speed and thus, supported the model where codon context affects the overall stacking energy that is obtained as each charged tRNA enters the ribosome's "A" site which ultimately affects the translational speed of the ribosome. Overall, these experiments suggest that codon context should be a variable considered when redesigning genes for heterologous expression (Chevance et al 2014). …”

Mg 2+ regulates transcription of mgtA in Salmonella Typhimurium via translation of proline codons during synthesis of the MgtL peptide

“…1). A stretch of three Pro residues could constitute a ribosome-stalling motif sequence (16)(17)(18)(19). Although mgtA-lacZ expression in the wildtype strain was up-regulated sevenfold upon decreasing [Mg 2+ ] from 1.6 mM (high) to 0.016 mM (low), the A86C mutant exhibited a 13-fold increased expression of the mgtA-lacZ fusion at high Mg 2+ , compared with the wild type (Fig.…”

Synonymous codons influencing gene expression in organisms

“…Apart from this, the association rate of ternary complex formation between an anticodon, the A-site of ribosome, and the mRNA may be dissimilar for different synonymous codons. 25 In addition, the impact of codon context during translation 26 and the effect of certain sequences in mRNA on ribosome movement during translation 27 are attributes of synonymous codons. Therefore, synonymous codons can influence gene expression at both the posttranscriptional and translational levels.…”