Qβ-Phage Resistance by Deletion of the Coiled-coil Motif in Elongation Factor Ts

Abstract

Elongation factor Ts (EF-Ts) is the guanine-nucleotide exchange factor of elongation factor Tu (EF-Tu), which promotes the binding of aminoacyl-tRNA to the mRNA-programmed ribosome in prokaryotes. The EF-Tu·EF-Ts complex, one of the EF-Tu complexes during protein synthesis, is also a component of RNA-dependent RNA polymerases like the polymerase from coli-phage Qβ. The present study shows that the Escherichia coli mutant GRd.tsf lacking the coiled-coil motif of EF-Ts is completely resistant to phage Qβ and that Qβ-polymerase complex formation is not observed. GRd.tsf is the first E. coli mutant ever described that is unable to form a Qβ-polymerase complex while still maintaining an almost normal growth behavior. The phage resistance correlates with an observed instability of the mutant EF-Tu·EF-Ts complex in the presence of guanine nucleotides. Thus, the mutant EF-Tu·EF-Ts is the first EF-Tu-EF·Ts complex ever described that is completely inactive in the Qβ-polymerase complex despite its almost full activity in protein synthesis. We propose that the role of EF-Ts in the Qβ-polymerase complex is to control and trap EF-Tu in a stable conformation with affinity for RNA templates while unable to bind aminoacyl-tRNA.