Human ribosomal protein S5 substitutes for its yeast homologue in vivo and determines binding of the hepatitis c virus (HCV) IRES element to the hybrid 40S ribosomes

Internal ribosome entry site (IRES) elements found in some viral and eukaryotic cellular mRNAs can recruit the small ribosomal subunit to the vicinity of initiation codon and initiate protein synthesis in a cap‐independent fashion. Hepatitis C virus (HCV) IRES is known to interact with the 40S ribosomal subunit alone, in the absence of any additional initiation factors or Met‐tRNAi. Ribosomal protein (rp) S5 has been suggested to be the prime candidate protein for binding of the HCV IRES to the 40S. Interestingly, HCV IRES has been shown to be able to bind purified human, but not yeast 40S. To test, whether the inability of HCV IRES to bind the yeast 40S ribosome, could be determined by the differences in sequence/structure between the yeast and human rpS5 proteins, we have substituted the yeast rpS5 with its human homologue. The mutant Saccharomyces cerevisiae strain bearing human ribosomal protein S5 instead of the yeast homologue was found to be viable and displayed only slightly reduced growth rates in comparison with the isogenic wt strain. Sucrose gradient centrifugation using purified hybrid yeast/human 40S ribosomes and in vitro transcribed HCV IRES element showed that hybrid 40S ribosomes in contrast to the wt ones did acquire the ability to bind the HCV IRES.