Nucleic Acids Research, 2003, Vol. 31, No. 6 1735-1743
© 2003 Oxford University Press
An NMR study on the interaction of Escherichia coli DinI with RecA–ssDNA complexes
1 Cellular and Molecular Biology Laboratory, RIKEN, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan, 2 The Graduate School of Science and Engineering, Saitama University, 255 Shimo-Okubo, Saitama-shi, Saitama 338-8570, Japan, 3 CREST/Japan Science and Technology Corporation (JST) 4-1-8 Kawaguchi Center Building, Honmachi, Kawaguchi-shi, Saitama 332-0012, Japan, 4 Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan, 5 Cellular Signaling Laboratory, RIKEN Harima Institute at SPring-8, 1-1-1 Kohto, Mikazuki-cho, Sayo, Hyogo 679-5143, Japan and 6 Protein Research Group, RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan
*To whom correspondence should be addressed at Cellular and Molecular Biology Laboratory, RIKEN, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan. Tel: +81 48 467 9537; Fax: +81 48 462 4671; Email: tshibata{at}postman.riken.go.jp
The SOS response, a set of cellular phenomena exhibited by eubacteria, is initiated by various causes that include DNA damage-induced replication arrest, and is positively regulated by the co- protease activity of RecA. Escherichia coli DinI, a LexA-regulated SOS gene product, shuts off the initiation of the SOS response when overexpressed in vivo. Biochemical and genetic studies indicated that DinI physically interacts with RecA to inhibit its co-protease activity. Using nuclear magnetic resonance (NMR) spectroscopy, we show that DinI tightly binds to the central region of RecA (between the N- and C-terminal domains) and that this interaction is enhanced upon the oligomerisation of RecA. On the other hand, DinI did not inhibit the interaction between 4mer single-stranded (ss)DNA and RecA– ATP
S, but had a slight effect on the structure of ssDNA–RecA–ATPS complexes involving 8mer and 12mer ssDNA. We hypothesise that prevention of repressor binding to the intermolecular cleft region of RecA protomers by DinI, with the possibility of a slight conformational change induced in the DinI-bound ssDNA–RecA–ATPS complex, together function to inhibit the co-protease activity of RecA.
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