Near-atomic structural model for bacterial DNA replication initiation complex and its functional insights

• Published in: Proceedings of the National Academy of Sciences - PNAS 2016-12, Vol.113 (50), p.E8021-E8030
• Main Authors: Shimizu, Masahiro, Noguchi, Yasunori, Sakiyama, Yukari, Kawakami, Hironori, Katayama, Tsutomu, Takada, Shoji
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - metabolism; Bacterial Proteins - chemistry; Bacterial Proteins - metabolism; Base Sequence; Biochemistry; Biological Sciences; Chromosomes; Computer Simulation; Deoxyribonucleic acid; DNA; DNA Replication - genetics; DNA, Bacterial - chemistry; DNA, Bacterial - genetics; DNA, Bacterial - metabolism; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - metabolism; E coli; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - metabolism; Integration Host Factors - chemistry; Integration Host Factors - metabolism; Models, Molecular; Molecular biology; Multiprotein Complexes - chemistry; Multiprotein Complexes - metabolism; Origin Recognition Complex - chemistry; Origin Recognition Complex - metabolism; Physical Sciences; PNAS Plus; Protein Interaction Domains and Motifs; Proteins
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1609649113

Upon DNA replication initiation in Escherichia coli, the initiator protein DnaA forms higher-order complexes with the chromosomal origin oriC and a DNA-bending protein IHF. Although tertiary structures of DnaA and IHF have previously been elucidated, dynamic structures of oriC–DnaA–IHF complexes remain unknown. Here, combining computer simulations with biochemical assays, we obtained models at almost-atomic resolution for the central part of the oriC–DnaA–IHF complex. This complex can be divided into three subcomplexes; the left and right subcomplexes include pentameric DnaA bound in a head-to-tail manner and the middle subcomplex contains only a single DnaA. In the left and right subcomplexes, DnaA ATPases associated with various cellular activities (AAA+) domain III formed helices with specific structural differences in interdomain orientations, provoking a bend in the bound DNA. In the left subcomplex a continuous DnaA chain exists, including insertion of IHF into the DNA looping, consistent with the DNA unwinding function of the complex. The intervening spaces in those subcomplexes are crucial for DNA unwinding and loading of DnaB helicases. Taken together, this model provides a reasonable near-atomic level structural solution of the initiation complex, including the dynamic conformations and spatial arrangements of DnaA subcomplexes.