title: Assembly of Helicobacter pylori initiation complex is determined by sequence-specific and topology-sensitive DnaA-oriC interactions.
abstract: In bacteria, chromosome replication is initiated by binding of the DnaA initiator protein to DnaA boxes located in the origin of chromosomal replication (oriC). This leads to DNA helix opening within the DNA-unwinding element. Helicobacter pylori oriC, the first bipartite origin identified in Gram-negative bacteria, contains two subregions, oriC1 and oriC2, flanking the dnaA gene. The DNA-unwinding element region is localized in the oriC2 subregion downstream of dnaA. Surprisingly, oriC2-DnaA interactions were shown to depend on DNA topology, which is unusual in bacteria but is similar to initiator-origin interactions observed in higher organisms. In this work, we identified three DnaA boxes in the oriC2 subregion, two of which were bound only as supercoiled DNA. We found that all three DnaA boxes play important roles in orisome assembly and subsequent DNA unwinding, but different functions can be assigned to individual boxes. This suggests that the H. pylori oriC may be functionally divided, similar to what was described recently for Escherichia coli oriC. On the basis of these results, we propose a model of initiation complex formation in H. pylori.
authors.affiliation: Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Department of Microbiology, Weigla 12, 53-114 Wrocław, Poland. Electronic address: rafal.donczew@iitd.pan.wroc.pl.
authors.forename: Rafał
authors.initials: R
authors.lastname: Donczew
authors.affiliation: Max Planck Institute for Molecular Genetics, Ihnestrasse 63-73, 14195 Berlin, Germany. Electronic address: mielke@molgen.mpg.de.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Mielke
authors.affiliation: Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Department of Microbiology, Weigla 12, 53-114 Wrocław, Poland. Electronic address: pawel.jaworski@iitd.pan.wroc.pl.
authors.forename: Paweł
authors.initials: P
authors.lastname: Jaworski
authors.affiliation: Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Department of Microbiology, Weigla 12, 53-114 Wrocław, Poland; University of Wrocław, Faculty of Biotechnology, Joliot-Curie 14a, 50-138 Wrocław, Poland. Electronic address: jolanta.zakrzewska@uni.wroc.pl.
authors.forename: Jolanta
authors.initials: J
authors.lastname: Zakrzewska-Czerwińska
authors.affiliation: Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Department of Microbiology, Weigla 12, 53-114 Wrocław, Poland. Electronic address: zawilak@iitd.pan.wroc.pl.
authors.forename: Anna
authors.initials: A
authors.lastname: Zawilak-Pawlik
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002876:Chromosomes, Bacterial / Q000235:genetics; D004261:DNA Replication; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D004278:DNA, Superhelical / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016480:Helicobacter pylori / Q000235:genetics / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D051738:Origin Recognition Complex / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; C114650:OriC chromosomal replication origin; D051738:Origin Recognition Complex
keywords: DNA replication; DNA sequence; DNA topology; oriC; orisome formation