title: The dimerization domain of R6K plasmid replication initiator protein pi revealed by analysis of a truncated protein.
abstract: Replication of plasmid R6K is controlled by the homodimeric initiator protein pi, which binds to seven 22-bp direct repeats (iterons) in the gamma-origin. One of the genetically engineered pi variants (delta C164 pi) contains only the 164 N-terminal amino acids (aa) of the 305-aa pi molecule. This truncated pi polypeptide retains the ability to function as a specific inhibitor of R6K replication in vivo, though it neither drives replication, nor binds to iterons [Greener et al., Mol. Gen. Genet. 224 (1990) 24-32]. In order to define the region of pi responsible for dimerization, we have performed chemical crosslinking experiments with purified delta C164 pi and shown that this polypeptide is dimeric. We did not observe an exchange between protein monomers upon mixing of wild-type pi and delta C164 pi homodimers. However, heterodimers, as well as each type of homodimers, were formed when these polypeptides refolded after guanidine hydrochloride treatment. Thus, both dimerization and dimer stability are determined by the N-terminal domain of pi. We speculate that these properties might depend on the leucine zipper and RGD motifs that have been identified in the two regions of the N-terminal domain of pi.
authors.affiliation: Department of Bacteriology, University of Wisconsin, Madison 53706.
authors.forename: I
authors.initials: I
authors.lastname: Levchenko
authors.forename: D
authors.initials: D
authors.lastname: York
authors.forename: M
authors.initials: M
authors.lastname: Filutowicz
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001704:Biopolymers; D004265:DNA Helicases; D004261:DNA Replication; D004268:DNA-Binding Proteins; D008969:Molecular Sequence Data; D017510:Protein Folding; D011815:R Factors; D015534:Trans-Activators
chemical_list: D001426:Bacterial Proteins; D001704:Biopolymers; D004268:DNA-Binding Proteins; D015534:Trans-Activators; C045264:replication initiator protein; D004265:DNA Helicases