Regulation of GTPases in the bacterial translation machinery

Several GTPases participate in bacterial protein biosynthesis. Initiation f actor 2 controls the formation of the ribosomal initiation complex and plac es initiator fMet-tRNA(fMet) in the ribosomal P-site, Elongation factors Tu and G are responsible for codon-specific binding of the aminoacyl-tRNA to the A-site, and peptidyl-tRNA to the P-site, respectively, during the elong ation phase of protein biosynthesis. Release factor 3, a GTPase which is no t ubiquitous, is involved in termination and release of the nascent polypep tide. Other translation factors, including initiation factors 1 and 3, elon gation factor Ts, release factors 1 and 2, and ribosomal release factor do not belong to the family of GTP/GDP binding proteins. The guanosine nucleot ide binding domains of the GTPases involved in translation are structurally related to the G alpha subunit Of heterotrimeric G proteins and to the pro teins of the Ras family, We have identified and sequenced all genes coding for translation factors i n the extreme thermophile Thermus thermophilus, The proteins were overprodu ced in Escherichia coli, purified, biochemically characterised and used for crystallisation and structural analysis. Further biochemical investigation s were aimed at gaining insight into the molecular mechanism underlying the regulation of the GTPase activity of the translation factors, and to eluci date the role of their ribosomal binding sites in this process.