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. 2000 Nov;9(11):2161–2169. doi: 10.1110/ps.9.11.2161

Solution structure of DinI provides insight into its mode of RecA inactivation.

B E Ramirez 1, O N Voloshin 1, R D Camerini-Otero 1, A Bax 1
PMCID: PMC2144493  PMID: 11152126

Abstract

The Escherichia coli RecA protein triggers both DNA repair and mutagenesis in a process known as the SOS response. The 81-residue E. coli protein DinI inhibits activity of RecA in vivo. The solution structure of DinI has been determined by multidimensional triple resonance NMR spectroscopy, using restraints derived from two sets of residual dipolar couplings, obtained in bicelle and phage media, supplemented with J couplings and a moderate number of NOE restraints. DinI has an alpha/beta fold comprised of a three-stranded beta-sheet and two alpha-helices. The beta-sheet topology is unusual: the central strand is flanked by a parallel and an antiparallel strand and the sheet is remarkably flat. The structure of DinI shows that six negatively charged Glu and Asp residues on DinI's kinked C-terminal alpha-helix form an extended, negatively charged ridge. We propose that this ridge mimics the electrostatic character of the DNA phospodiester backbone, thereby enabling DinI to compete with single-stranded DNA for RecA binding. Biochemical data confirm that DinI is able to displace ssDNA from RecA.

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Selected References

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  1. Clore G. M., Brünger A. T., Karplus M., Gronenborn A. M. Application of molecular dynamics with interproton distance restraints to three-dimensional protein structure determination. A model study of crambin. J Mol Biol. 1986 Oct 5;191(3):523–551. doi: 10.1016/0022-2836(86)90146-4. [DOI] [PubMed] [Google Scholar]
  2. Clore G. M., Gronenborn A. M., Bax A. A robust method for determining the magnitude of the fully asymmetric alignment tensor of oriented macromolecules in the absence of structural information. J Magn Reson. 1998 Jul;133(1):216–221. doi: 10.1006/jmre.1998.1419. [DOI] [PubMed] [Google Scholar]
  3. Cornilescu G., Delaglio F., Bax A. Protein backbone angle restraints from searching a database for chemical shift and sequence homology. J Biomol NMR. 1999 Mar;13(3):289–302. doi: 10.1023/a:1008392405740. [DOI] [PubMed] [Google Scholar]
  4. Cox M. M. Recombinational DNA repair in bacteria and the RecA protein. Prog Nucleic Acid Res Mol Biol. 1999;63:311–366. doi: 10.1016/s0079-6603(08)60726-6. [DOI] [PubMed] [Google Scholar]
  5. Delaglio F., Grzesiek S., Vuister G. W., Zhu G., Pfeifer J., Bax A. NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J Biomol NMR. 1995 Nov;6(3):277–293. doi: 10.1007/BF00197809. [DOI] [PubMed] [Google Scholar]
  6. Drohat A. C., Tjandra N., Baldisseri D. M., Weber D. J. The use of dipolar couplings for determining the solution structure of rat apo-S100B(betabeta). Protein Sci. 1999 Apr;8(4):800–809. doi: 10.1110/ps.8.4.800. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gehring K., Ekiel I. H(C)CH-COSY and (H)CCH-COSY experiments for 13C-labeled proteins in H2O solution. J Magn Reson. 1998 Nov;135(1):185–193. doi: 10.1006/jmre.1998.1543. [DOI] [PubMed] [Google Scholar]
  8. Grzesiek S., Bax A., Hu J. S., Kaufman J., Palmer I., Stahl S. J., Tjandra N., Wingfield P. T. Refined solution structure and backbone dynamics of HIV-1 Nef. Protein Sci. 1997 Jun;6(6):1248–1263. doi: 10.1002/pro.5560060613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Grzesiek S., Vuister G. W., Bax A. A simple and sensitive experiment for measurement of JCC couplings between backbone carbonyl and methyl carbons in isotopically enriched proteins. J Biomol NMR. 1993 Jul;3(4):487–493. doi: 10.1007/BF00176014. [DOI] [PubMed] [Google Scholar]
  10. Güntert P., Braun W., Wüthrich K. Efficient computation of three-dimensional protein structures in solution from nuclear magnetic resonance data using the program DIANA and the supporting programs CALIBA, HABAS and GLOMSA. J Mol Biol. 1991 Feb 5;217(3):517–530. doi: 10.1016/0022-2836(91)90754-t. [DOI] [PubMed] [Google Scholar]
  11. Hansen M. R., Mueller L., Pardi A. Tunable alignment of macromolecules by filamentous phage yields dipolar coupling interactions. Nat Struct Biol. 1998 Dec;5(12):1065–1074. doi: 10.1038/4176. [DOI] [PubMed] [Google Scholar]
  12. Holm L., Sander C. Protein structure comparison by alignment of distance matrices. J Mol Biol. 1993 Sep 5;233(1):123–138. doi: 10.1006/jmbi.1993.1489. [DOI] [PubMed] [Google Scholar]
  13. Koradi R., Billeter M., Wüthrich K. MOLMOL: a program for display and analysis of macromolecular structures. J Mol Graph. 1996 Feb;14(1):51-5, 29-32. doi: 10.1016/0263-7855(96)00009-4. [DOI] [PubMed] [Google Scholar]
  14. Kuszewski J., Gronenborn A. M., Clore G. M. Improvements and extensions in the conformational database potential for the refinement of NMR and X-ray structures of proteins and nucleic acids. J Magn Reson. 1997 Mar;125(1):171–177. doi: 10.1006/jmre.1997.1116. [DOI] [PubMed] [Google Scholar]
  15. Liu D., Ishima R., Tong K. I., Bagby S., Kokubo T., Muhandiram D. R., Kay L. E., Nakatani Y., Ikura M. Solution structure of a TBP-TAF(II)230 complex: protein mimicry of the minor groove surface of the TATA box unwound by TBP. Cell. 1998 Sep 4;94(5):573–583. doi: 10.1016/s0092-8674(00)81599-8. [DOI] [PubMed] [Google Scholar]
  16. Nicholls A., Sharp K. A., Honig B. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins. 1991;11(4):281–296. doi: 10.1002/prot.340110407. [DOI] [PubMed] [Google Scholar]
  17. Ottiger M., Bax A. Characterization of magnetically oriented phospholipid micelles for measurement of dipolar couplings in macromolecules. J Biomol NMR. 1998 Oct;12(3):361–372. doi: 10.1023/a:1008366116644. [DOI] [PubMed] [Google Scholar]
  18. Ottiger M., Delaglio F., Bax A. Measurement of J and dipolar couplings from simplified two-dimensional NMR spectra. J Magn Reson. 1998 Apr;131(2):373–378. doi: 10.1006/jmre.1998.1361. [DOI] [PubMed] [Google Scholar]
  19. Sanders C. R., 2nd, Schwonek J. P. Characterization of magnetically orientable bilayers in mixtures of dihexanoylphosphatidylcholine and dimyristoylphosphatidylcholine by solid-state NMR. Biochemistry. 1992 Sep 22;31(37):8898–8905. doi: 10.1021/bi00152a029. [DOI] [PubMed] [Google Scholar]
  20. Selmer M., Al-Karadaghi S., Hirokawa G., Kaji A., Liljas A. Crystal structure of Thermotoga maritima ribosome recycling factor: a tRNA mimic. Science. 1999 Dec 17;286(5448):2349–2352. doi: 10.1126/science.286.5448.2349. [DOI] [PubMed] [Google Scholar]
  21. Shinagawa H., Iwasaki H., Kato T., Nakata A. RecA protein-dependent cleavage of UmuD protein and SOS mutagenesis. Proc Natl Acad Sci U S A. 1988 Mar;85(6):1806–1810. doi: 10.1073/pnas.85.6.1806. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Struppe J., Vold R. R. Dilute bicellar solutions for structural NMR work. J Magn Reson. 1998 Dec;135(2):541–546. doi: 10.1006/jmre.1998.1605. [DOI] [PubMed] [Google Scholar]
  23. Subramanya H. S., Roper D. I., Dauter Z., Dodson E. J., Davies G. J., Wilson K. S., Wigley D. B. Enzymatic ketonization of 2-hydroxymuconate: specificity and mechanism investigated by the crystal structures of two isomerases. Biochemistry. 1996 Jan 23;35(3):792–802. doi: 10.1021/bi951732k. [DOI] [PubMed] [Google Scholar]
  24. Sugimoto H., Taniguchi M., Nakagawa A., Tanaka I., Suzuki M., Nishihira J. Crystal structure of human D-dopachrome tautomerase, a homologue of macrophage migration inhibitory factor, at 1.54 A resolution. Biochemistry. 1999 Mar 16;38(11):3268–3279. doi: 10.1021/bi982184o. [DOI] [PubMed] [Google Scholar]
  25. Tjandra N., Bax A. Direct measurement of distances and angles in biomolecules by NMR in a dilute liquid crystalline medium. Science. 1997 Nov 7;278(5340):1111–1114. doi: 10.1126/science.278.5340.1111. [DOI] [PubMed] [Google Scholar]
  26. Tjandra N., Omichinski J. G., Gronenborn A. M., Clore G. M., Bax A. Use of dipolar 1H-15N and 1H-13C couplings in the structure determination of magnetically oriented macromolecules in solution. Nat Struct Biol. 1997 Sep;4(9):732–738. doi: 10.1038/nsb0997-732. [DOI] [PubMed] [Google Scholar]
  27. Vijay-Kumar S., Bugg C. E., Cook W. J. Structure of ubiquitin refined at 1.8 A resolution. J Mol Biol. 1987 Apr 5;194(3):531–544. doi: 10.1016/0022-2836(87)90679-6. [DOI] [PubMed] [Google Scholar]
  28. Vold R. R., Prosser R. S., Deese A. J. Isotropic solutions of phospholipid bicelles: a new membrane mimetic for high-resolution NMR studies of polypeptides. J Biomol NMR. 1997 Apr;9(3):329–335. doi: 10.1023/a:1018643312309. [DOI] [PubMed] [Google Scholar]
  29. Walker G. C. Skiing the black diamond slope: progress on the biochemistry of translesion DNA synthesis. Proc Natl Acad Sci U S A. 1998 Sep 1;95(18):10348–10350. doi: 10.1073/pnas.95.18.10348. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Yasuda T., Morimatsu K., Horii T., Nagata T., Ohmori H. Inhibition of Escherichia coli RecA coprotease activities by DinI. EMBO J. 1998 Jun 1;17(11):3207–3216. doi: 10.1093/emboj/17.11.3207. [DOI] [PMC free article] [PubMed] [Google Scholar]

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