title: The N-terminus promotes oligomerization of the Escherichia coli initiator protein DnaA.
abstract: Initiation of chromosome replication in Escherichia coli is governed by the interaction of the initiator protein DnaA with the replication origin oriC. Here we present evidence that homo-oligomerization of DnaA via its N-terminus (amino acid residues 1-86) is also essential for initiation. Results from solid-phase protein-binding assays indicate that residues 1-86 (or 1-77) of DnaA are necessary and sufficient for self interaction. Using a 'one-hybrid-system' we found that the DnaA N-terminus can functionally replace the dimerization domain of coliphage lambda cl repressor: a lambdacl-DnaA chimeric protein inhibits lambda plasmid replication as efficiently as lambdacI repressor. DnaA derivatives with deletions in the N-terminus are incapable of supporting chromosome replication from oriC, and, conversely, overexpression of the DnaA N-terminus inhibits initiation in vivo. Together, these results indicate that (i) oligomerization of DnaA N-termini is essential for protein function during initiation, and (ii) oligomerization does not require intramolecular cross-talk with the nucleotide-binding domain III or the DNA-binding domain IV. We propose that E. coli DnaA is composed of largely independent domains - or modules - each contributing a partial, though essential, function to the proper functioning of the 'holoprotein'.
authors.affiliation: Max-Planck-Institut für Molekulare Genetik, Ihnestrasse 73, D-14195 Berlin-Dahlem, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Weigel
authors.forename: A
authors.initials: A
authors.lastname: Schmidt
authors.forename: H
authors.initials: H
authors.lastname: Seitz
authors.forename: D
authors.initials: D
authors.lastname: Tüngler
authors.forename: M
authors.initials: M
authors.lastname: Welzeck
authors.forename: W
authors.initials: W
authors.lastname: Messer
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D009698:Nucleoproteins / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D018741:Replication Origin; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; D009698:Nucleoproteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D012097:Repressor Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins