title: Dimerization of the RelA protein of Escherichia coli.
abstract: The RelA protein of Escherichia coli is a ribosome-associated (p)ppGpp synthetase that is activated by amino acid deprivation. It was recently reported that the activity of RelA is regulated by oligomerization mediated by the C-terminal domain of RelA. The oligomerization of RelA is further characterized in this study. The C-terminal domain consisting of amino acids 455-744, designated 'RelA, formed homooligomers as well as heterooligomers with RelA as demonstrated by copurification of RelA and 'RelA and by an affinity blotting assay. Glutaraldehyde-induced cross-linking indicated that the oligomer was a dimer. The functional analysis of 'RelA was based on a combination of yeast two-hybrid analysis, the determination of the effects of overexpression of 'RelA derivatives on the stringent response, and the cellular localization of the overexpressed 'RelA derivatives. These studies indicated that two regions, designated 'RelA-1 (amino acids 455-538) and 'RelA-2 (amino acids 550-682), were involved in dimerization. The involvement of one of these two regions, RelA-2, is consistent with a previous site-directed mutagenesis study. In addition to dimerization, 'RelA-2 apparently contained the main ribosome-binding domain of RelA. The third region, 'RelA-3 (amino acids 682-744), was not involved in either dimerization or ribosome binding. The overexpression of 'RelA-1 and 'RelA-2, but not 'RelA-3, inhibited the stringent response. These results support the previously proposed model which suggests a role for oligomerization in the regulation of (p)ppGpp synthetase.
authors.affiliation: Department of Biochemistry and Microbiology, University of Victoria, BC, Canada.
authors.forename: X
authors.initials: X
authors.lastname: Yang
authors.forename: E E
authors.initials: EE
authors.lastname: Ishiguro
mesh_terms: D000345:Affinity Labels; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism*; D008025:Ligases / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
chemical_list: D000345:Affinity Labels; D008025:Ligases; C028812:guanosine 3',5'-polyphosphate synthetases