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Protein–nucleotide interactions in E. coli DNA topoisomerase I

Nature Structural Biology volume 6, pages 961–968 (1999)Cite this article

Abstract

DNA topoisomerases are the enzymes responsible for controlling and maintaining the topological states of DNA. Type IA enzymes work by transiently breaking the phosphodiester backbone of one strand to allow passage of another strand through the break. The protein has to perform complex rearrangements of the DNA, and hence it is likely that different regions of the enzyme bind DNA with different affinities. In order to identify some of the DNA binding sites in the protein, we have solved the structures of several complexes of the 67 kDa N-terminal fragment of Escherichia coli DNA topoisomerase I with mono- and trinucleotides. There are five different binding sites in the complexes, one of which is adjacent to the active site. Two other sites are in the central hole of the protein and may represent general DNA binding regions. The positions of these sites allow us to identify different DNA binding regions and to understand their possible roles in the catalytic cycle.

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Figure 1: Overall structure of the 67 kDa fragment of E.coli DNA topoisomerase I showing the four domains and the locations of the five nucleotide binding sites.
Figure 2: Electron density stereo maps illustrating different nucleotide binding sites.
Figure 3: Models showing different conformations of the loop region in domain III.
Figure 4: Stereo view of the active site region.
Figure 5: Surface representations of a model for the open conformation of the 67 kDa fragment of E.coli DNA topoisomerase I.

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References

  1. Wang, J.C. DNA Topoisomerases. Annu. Rev. Biochem. 65, 635–692 (1996).

    Article  CAS  Google Scholar 

  2. Lindsley, J.E. & Wang, J.C. On the coupling between ATP usage and DNA transport by yeast DNA topoisomerase II. J. Biol. Chem. 268, 8096–8104 ( 1993).

    CAS  PubMed  Google Scholar 

  3. Caron, P.R. & Wang, J.C. Alignment of Primary Sequences of DNA Topoisomerases. Adv. Pharmacol. 29B 271 –297 (1994).

    Article  CAS  Google Scholar 

  4. Cheng, C., Kussie, P., Pavletich, N. & Shuman, S. Conservation of structure and mechanism between eukaryotic topoisomerase I and site-specific recombinases. Cell 92, 841–850 (1998).

    Article  CAS  Google Scholar 

  5. Sharma, A., Hanai, R. & Mondragón, A. Crystal structure of the N-terminal fragment of vaccinia virus DNA topoisomerase I at 1.6 A resolution. Structure 2, 767–777 (1994).

    Article  CAS  Google Scholar 

  6. Redinbo, M.R., Stewart, L., Kuhn, P., Champoux, J.J. & Hol, W.G. Crystal structures of human topoisomerase I in covalent and noncovalent complexes with DNA. Science 279, 1504–1513 (1998).

    Article  CAS  Google Scholar 

  7. Stewart, L., Redinbo, M.R., Qiu, X., Hol, W.G. & Champoux, J.J. A model for the mechanism of human topoisomerase I. Science 279, 1534–1541 (1998).

    Article  CAS  Google Scholar 

  8. Lima, C.D., Wang, J.C. & Mondragón, A. Three dimensional structure of the 67K N-terminal fragment of E. coli DNA topoisomerase I. Nature 367, 138–146 (1994).

    Article  CAS  Google Scholar 

  9. Yu, L., Zhu, C.X., Tse-Dinh, Y.C. & Fesik, S.W. Solution structure of the Cterminal single-stranded DNA-binding domain of Escherichia coli topoisomerase I. Biochemistry 34, 7622–7628 (1995).

    Article  CAS  Google Scholar 

  10. Feinberg, H., Lima, C.D. & Mondragón, A. Conformational changes in E. coli DNA topoisomerase I. Nature Struct. Biol. 6, 918– 922(1999).

    Article  CAS  Google Scholar 

  11. Kirkegaard, K. & Wang, J.C. Bacterial DNA topoisomerase I can relax positively supercoiled DNA containing a single stranded loop. J. Mol. Biol. 185, 625– 637 (1985).

    Article  CAS  Google Scholar 

  12. Kirkegaard, K. & Wang, J.C. Escherichia coli DNA topoisomerase I catalyzed linking of single-stranded rings of complementary base sequences. Nucleic Acids Res. 5, 3811 –3820 (1978).

    Article  CAS  Google Scholar 

  13. Tse, Y.C., Kirkegaard, K. & Wang, J.C. Covalent bonds between protein and DNA. J. Biol. Chem. 255, 5560–5565 (1980).

    CAS  PubMed  Google Scholar 

  14. Zhang, H.L., Malpure, S. & DiGate, R.J. Escherichia coli DNA topoisomerase III is a site-specific DNA binding protein that binds asymmetrically to its cleavage site. J. Biol. Chem. 270, 23700– 23705 (1995).

    Article  CAS  Google Scholar 

  15. Beran-Steed, R.K. & Tse-Dinh, Y.C. The carboxy terminal domain of Escherichia coli DNA topoisomerase I confers higher affinity to DNA. Proteins 6, 249– 258 (1989).

    Article  CAS  Google Scholar 

  16. Tse-Dinh, Y.C. & Beran-Seed, R.K. Escherichia coli DNA topoisomerase I is a zinc metalloprotein with three repetitive zinc binding domains. J. Biol. Chem. 263, 15857–15859 (1988).

    CAS  PubMed  Google Scholar 

  17. Zhang, H.L. & DiGate, R.J. The C-terminal residues of Escherichia coli DNA topoisomerase III are involved in substrate binding. J. Biol. Chem. 269, 9052– 9059 (1994).

    CAS  PubMed  Google Scholar 

  18. Zhang, H.L., Malpure, S., Li, Z., Hiasa, H. & DiGate, R.J. The role of the C-terminal amino acid residues in Escherichia coli DNA topoisomerase III-mediated catalysis. J. Biol. Chem. 271, 9039–9045 (1996).

    Article  CAS  Google Scholar 

  19. Domanico, P.L. & Tse-Dinh, Y.C. Cleavage of dT8 and dT8 Phosphorothyioyl Analogues by Escherichia coli DNA Topoisomerase I: Product and Rate Analysis. Biochemistry 27, 6365–6371 (1988).

    Article  CAS  Google Scholar 

  20. Wolfenden, R., Ridgway, C. & Young, G. Spontaneous hydrolysis of ionized phosphate monoesters and diesters and the proficiencies of phosphatases and phosphodiesterases as catalysts. J. Am. Chem. Soc. 120, 833 –834 (1998).

    Article  CAS  Google Scholar 

  21. Ding, J., Koellner, G., Grunert, H.P. & Saenger, W. Crystal structure of ribonuclease T1 complexed with adenosine 2'-monophosphate at 1.8 A resolution. J. Biol. Chem. 266, 15128–15134 (1991).

    CAS  PubMed  Google Scholar 

  22. Lima, C.D., Wang, J.C. & Mondragón, A. Crystallization of a 67 kDa fragment of Escherichia coli DNA topoisomerase I. J. Mol. Biol. 232, 1213–1216 (1993).

    Article  CAS  Google Scholar 

  23. Otwinowski, Z. & Minor, W. Processing of X-ray Diffraction Data Collected in Oscillation Mode. Methods Enzymol. 276A 307–326 ( 1997).

    Article  Google Scholar 

  24. Collaborative Computational Project 4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D50, 760–763 ( 1994).

  25. Brunger, A.T. X-PLOR, Version 3.1, a system for X-ray crystallography and NMR (Yale University Press, New Haven, Connecticut, 1992).

    Google Scholar 

  26. Jones, T.A., Zou, J.Y., Cowan, S.W. & Kjeldgaard, M. Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A47, 110–119 (1991).

    Article  CAS  Google Scholar 

  27. Kleywegt, G.J. & Jones, T.A. Databases in protein crystallography. Acta Crystallogr. D54, 1119–1131 (1998).

    CAS  Google Scholar 

  28. Branden, C.I. Relation between structure and function of alpha/beta proteins. Q. Rev. Biophys. 13, 317–338 (1980).

    Article  Google Scholar 

  29. Berger, J.M., Fass, D., Wang, J.C. & Harrison, S.C. Structural similarities between topoisomerases that cleave one or both DNA strands. Proc. Natl. Acad. Sci. USA 95, 7876– 81 (1998).

    Article  CAS  Google Scholar 

  30. Kraulis, P.J. MOLSCRIPT: a program to produce both detailed and schematic plots of protein structures. J. Appl. Crystallogr. 24, 946 –950 (1991).

    Article  Google Scholar 

  31. Evans, S.V. SETOR: Hardware lighted three dimensional solid model representations of macromolecules. J. Mol. Graphics 11, 134– 138 (1993).

    Article  CAS  Google Scholar 

  32. Bacon, D. & Anderson, W.F. A fast algorithm for rendering space-filling molecule pictures. J. Mol. Graph. 6, 219–220 (1988).

    Article  Google Scholar 

  33. Merritt, E.A. & Murphy, M.E.P. Raster3D version 2.0. A program for photorealistic molecular graphics. Acta Crystallogr. D50, 869–873 (1994).

    CAS  Google Scholar 

  34. Nicholls, A., Sharp, K.A. & Honig, B.H. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins 11, 281–286 ( 1991).

    Article  CAS  Google Scholar 

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Acknowledgements

We acknowledge the contributions of C.D. Lima and A. Patera to the early stages of this project. We thank past and present members of the laboratory for help with data collection and for comments and suggestions. We also thank R. DiGate, L. Godley, M. Gwynn, T. Jardetzky, K. Perry, X. Qiu, A. Rosenzweig, A. Tackle, J. Widom, X. Yang and members of the Center of Structural Biology for comments and suggestions. We thank BNLS, CHESS, DND-CAT, and SSRL for access to their beamlines and help during data collection. We thank M. Blum of MAR USA for lending us the MAR CCD detector used at SSRL. This work was supported by the NIH (A.M.) and by SmithKline Beecham Pharmaceuticals. Portions of this work were performed at the DuPont-Northwestern-Dow Collaborative Access Team (DND-CAT) Synchrotron Research Center at the Advanced Photon Source, supported by DuPont, Dow, NSF and the State of Illinois. Use of the APS was supported by the DOE. Portions of this work were performed at SSRL, which is operated by the DOE. The SSRL Biotechnology Program is supported by the NIH and the DOE.

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  1. Hadar Feinberg

    Present address: Department of Structural Biology, Stanford University School of Medicine, Fairchild Building, Stanford, California, 94305-5400, USA

Authors and Affiliations

  1. Department of Biochemistry, Molecular Biology and Cell Biology, Northwerstern University, 2153 Sheridan Road, Evanston, 60208, Illinois, USA

    Hadar Feinberg, Anita Changela & Alfonso Mondragón

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Correspondence to Alfonso Mondragón.

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Feinberg, H., Changela, A. & Mondragón, A. Protein–nucleotide interactions in E. coli DNA topoisomerase I. Nat Struct Mol Biol 6, 961–968 (1999). https://doi.org/10.1038/13333

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