Interchanging functionality among homologous elongation factors using signatures of heterotachy.
Numerous models of molecular evolution have been formulated to describe the forces that shape sequence divergence among homologous proteins. These models have greatly enhanced our understanding of evolutionary processes. Rarely are such models empirically tested in the laboratory, and even more rare, are such models exploited to generate novel molecules useful for synthetic biology. Here, we experimentally demonstrate that the heterotachy model of evolution captures signatures of functional divergence among homologous elongation factors (EFs) between bacterial EF-Tu and eukaryotic eEF1A. These EFs are GTPases that participate in protein translation by presenting aminoacylated-tRNAs to the ribosome. Upon release from the ribosome, the EFs are recharged by nucleotide exchange factors EF-Ts in bacteria or eEF1B in eukaryotes. The two nucleotide exchange factors perform analogous functions despite not being homologous proteins. The heterotachy model was used to identify a set of sites in eEF1A/EF-Tu associated with eEF1B binding in eukaryotes and another reciprocal set associated with EF-Ts binding in bacteria. Introduction of bacterial EF-Tu residues at these sites into eEF1A protein efficiently disrupted binding of cognate eEF1B as well as endowed eEF1A with the novel ability to bind bacterial EF-Ts. We further demonstrate that eEF1A variants, unlike yeast wild-type, can function in a reconstituted in vitro bacterial translation system.
School of Biology, Georgia Institute of Technology, Atlanta, GA, USA.
Ercan
E
Cacan
James T
JT
Kratzer
Megan F
MF
Cole
Eric A
EA
Gaucher
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D009711:Nucleotides / Q000235:genetics / Q000378:metabolism; D010444:Peptide Elongation Factor Tu / Q000235:genetics / Q000378:metabolism; D010445:Peptide Elongation Factors / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D058849:Protein Refolding; D012343:RNA, Transfer / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D009711:Nucleotides; D010445:Peptide Elongation Factors; C023512:elongation factor Ts; D012343:RNA, Transfer; D010444:Peptide Elongation Factor Tu