Biochemical characterization and functional analysis of CgtA, a monomeric GTP -binding protein from <italic>Caulobacter crescentus</italic>.

Abstract

The <italic>Caulobacter crescentus</italic> CgtA protein is a member of the essential Obg GTPase subfamily. The bacterial Obg proteins are widespread in different organisms and are likely to play critical yet unknown cellular roles. In this thesis project, to study the cellular function of CgtA, <italic> in vitro</italic> biochemical and kinetic parameters of GTP/GDP binding, exchange, and GTP hydrolysis at physiologically relevant conditions were characterized. The N-terminal half non-GTPase domain does not play a role in guanine nucleotide binding and exchange. Conserved amino acid residues in the switch-I domain were changed, and the corresponding mutants were examined for nucleotide binding, exchange, GTPase activity, as well as in vivo functioning. The association of CgtA with ribosome or ribosomal subunits, and consequently the effects of CgtA depletion on polyribosome profiles of <italic>Caulobacter</italic> were examined. Finally, a summary of research results and discussion of future direction is presented.