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. 1997 Oct 1;16(19):6044–6054. doi: 10.1093/emboj/16.19.6044

Crystal structure of the specific DNA-binding domain of Tc3 transposase of C.elegans in complex with transposon DNA.

G van Pouderoyen 1, R F Ketting 1, A Perrakis 1, R H Plasterk 1, T K Sixma 1
PMCID: PMC1170234  PMID: 9312061

Abstract

The crystal structure of the complex between the N-terminal DNA-binding domain of Tc3 transposase and an oligomer of transposon DNA has been determined. The specific DNA-binding domain contains three alpha-helices, of which two form a helix-turn-helix (HTH) motif. The recognition of transposon DNA by the transposase is mediated through base-specific contacts and complementarity between protein and sequence-dependent deformations of the DNA. The HTH motif makes four base-specific contacts with the major groove, and the N-terminus makes three base-specific contacts with the minor groove. The DNA oligomer adopts a non-linear B-DNA conformation, made possible by a stretch of seven G:C base pairs at one end and a TATA sequence towards the other end. Extensive contacts (seven salt bridges and 16 hydrogen bonds) of the protein with the DNA backbone allow the protein to probe and recognize the sequence-dependent DNA deformation. The DNA-binding domain forms a dimer in the crystals. Each monomer binds a separate transposon end, implying that the dimer plays a role in synapsis, necessary for the simultaneous cleavage of both transposon termini.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aldaz H., Schuster E., Baker T. A. The interwoven architecture of the Mu transposase couples DNA synapsis to catalysis. Cell. 1996 Apr 19;85(2):257–269. doi: 10.1016/s0092-8674(00)81102-2. [DOI] [PubMed] [Google Scholar]
  2. Ariyoshi M., Vassylyev D. G., Iwasaki H., Nakamura H., Shinagawa H., Morikawa K. Atomic structure of the RuvC resolvase: a holliday junction-specific endonuclease from E. coli. Cell. 1994 Sep 23;78(6):1063–1072. doi: 10.1016/0092-8674(94)90280-1. [DOI] [PubMed] [Google Scholar]
  3. Baker T. A., Luo L. Identification of residues in the Mu transposase essential for catalysis. Proc Natl Acad Sci U S A. 1994 Jul 5;91(14):6654–6658. doi: 10.1073/pnas.91.14.6654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Barlow D. J., Thornton J. M. Ion-pairs in proteins. J Mol Biol. 1983 Aug 25;168(4):867–885. doi: 10.1016/s0022-2836(83)80079-5. [DOI] [PubMed] [Google Scholar]
  5. Beamer L. J., Pabo C. O. Refined 1.8 A crystal structure of the lambda repressor-operator complex. J Mol Biol. 1992 Sep 5;227(1):177–196. doi: 10.1016/0022-2836(92)90690-l. [DOI] [PubMed] [Google Scholar]
  6. Cai M., Zheng R., Caffrey M., Craigie R., Clore G. M., Gronenborn A. M. Solution structure of the N-terminal zinc binding domain of HIV-1 integrase. Nat Struct Biol. 1997 Jul;4(7):567–577. doi: 10.1038/nsb0797-567. [DOI] [PubMed] [Google Scholar]
  7. Chaconas G., Lavoie B. D., Watson M. A. DNA transposition: jumping gene machine, some assembly required. Curr Biol. 1996 Jul 1;6(7):817–820. doi: 10.1016/s0960-9822(02)00603-6. [DOI] [PubMed] [Google Scholar]
  8. Colloms S. D., van Luenen H. G., Plasterk R. H. DNA binding activities of the Caenorhabditis elegans Tc3 transposase. Nucleic Acids Res. 1994 Dec 25;22(25):5548–5554. doi: 10.1093/nar/22.25.5548. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Czerny T., Schaffner G., Busslinger M. DNA sequence recognition by Pax proteins: bipartite structure of the paired domain and its binding site. Genes Dev. 1993 Oct;7(10):2048–2061. doi: 10.1101/gad.7.10.2048. [DOI] [PubMed] [Google Scholar]
  10. Doak T. G., Doerder F. P., Jahn C. L., Herrick G. A proposed superfamily of transposase genes: transposon-like elements in ciliated protozoa and a common "D35E" motif. Proc Natl Acad Sci U S A. 1994 Feb 1;91(3):942–946. doi: 10.1073/pnas.91.3.942. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Doolittle R. F., Feng D. F., Johnson M. S., McClure M. A. Origins and evolutionary relationships of retroviruses. Q Rev Biol. 1989 Mar;64(1):1–30. doi: 10.1086/416128. [DOI] [PubMed] [Google Scholar]
  12. Dyda F., Hickman A. B., Jenkins T. M., Engelman A., Craigie R., Davies D. R. Crystal structure of the catalytic domain of HIV-1 integrase: similarity to other polynucleotidyl transferases. Science. 1994 Dec 23;266(5193):1981–1986. doi: 10.1126/science.7801124. [DOI] [PubMed] [Google Scholar]
  13. Epstein J. A., Glaser T., Cai J., Jepeal L., Walton D. S., Maas R. L. Two independent and interactive DNA-binding subdomains of the Pax6 paired domain are regulated by alternative splicing. Genes Dev. 1994 Sep 1;8(17):2022–2034. doi: 10.1101/gad.8.17.2022. [DOI] [PubMed] [Google Scholar]
  14. Epstein J., Cai J., Glaser T., Jepeal L., Maas R. Identification of a Pax paired domain recognition sequence and evidence for DNA-dependent conformational changes. J Biol Chem. 1994 Mar 18;269(11):8355–8361. [PubMed] [Google Scholar]
  15. Fayet O., Ramond P., Polard P., Prère M. F., Chandler M. Functional similarities between retroviruses and the IS3 family of bacterial insertion sequences? Mol Microbiol. 1990 Oct;4(10):1771–1777. doi: 10.1111/j.1365-2958.1990.tb00555.x. [DOI] [PubMed] [Google Scholar]
  16. Feng J. A., Johnson R. C., Dickerson R. E. Hin recombinase bound to DNA: the origin of specificity in major and minor groove interactions. Science. 1994 Jan 21;263(5145):348–355. doi: 10.1126/science.8278807. [DOI] [PubMed] [Google Scholar]
  17. Franz G., Loukeris T. G., Dialektaki G., Thompson C. R., Savakis C. Mobile Minos elements from Drosophila hydei encode a two-exon transposase with similarity to the paired DNA-binding domain. Proc Natl Acad Sci U S A. 1994 May 24;91(11):4746–4750. doi: 10.1073/pnas.91.11.4746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hol W. G., van Duijnen P. T., Berendsen H. J. The alpha-helix dipole and the properties of proteins. Nature. 1978 Jun 8;273(5662):443–446. doi: 10.1038/273443a0. [DOI] [PubMed] [Google Scholar]
  19. 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]
  20. Ivics Z., Izsvak Z., Minter A., Hackett P. B. Identification of functional domains and evolution of Tc1-like transposable elements. Proc Natl Acad Sci U S A. 1996 May 14;93(10):5008–5013. doi: 10.1073/pnas.93.10.5008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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 A. 1991 Mar 1;47(Pt 2):110–119. doi: 10.1107/s0108767390010224. [DOI] [PubMed] [Google Scholar]
  22. Joshua-Tor L., Frolow F., Appella E., Hope H., Rabinovich D., Sussman J. L. Three-dimensional structures of bulge-containing DNA fragments. J Mol Biol. 1992 May 20;225(2):397–431. doi: 10.1016/0022-2836(92)90929-e. [DOI] [PubMed] [Google Scholar]
  23. Katayanagi K., Miyagawa M., Matsushima M., Ishikawa M., Kanaya S., Ikehara M., Matsuzaki T., Morikawa K. Three-dimensional structure of ribonuclease H from E. coli. Nature. 1990 Sep 20;347(6290):306–309. doi: 10.1038/347306a0. [DOI] [PubMed] [Google Scholar]
  24. Katayanagi K., Miyagawa M., Matsushima M., Ishikawa M., Kanaya S., Nakamura H., Ikehara M., Matsuzaki T., Morikawa K. Structural details of ribonuclease H from Escherichia coli as refined to an atomic resolution. J Mol Biol. 1992 Feb 20;223(4):1029–1052. doi: 10.1016/0022-2836(92)90260-q. [DOI] [PubMed] [Google Scholar]
  25. Kissinger C. R., Liu B. S., Martin-Blanco E., Kornberg T. B., Pabo C. O. Crystal structure of an engrailed homeodomain-DNA complex at 2.8 A resolution: a framework for understanding homeodomain-DNA interactions. Cell. 1990 Nov 2;63(3):579–590. doi: 10.1016/0092-8674(90)90453-l. [DOI] [PubMed] [Google Scholar]
  26. Klemm J. D., Rould M. A., Aurora R., Herr W., Pabo C. O. Crystal structure of the Oct-1 POU domain bound to an octamer site: DNA recognition with tethered DNA-binding modules. Cell. 1994 Apr 8;77(1):21–32. doi: 10.1016/0092-8674(94)90231-3. [DOI] [PubMed] [Google Scholar]
  27. Kleywegt G. J., Jones T. A. Efficient rebuilding of protein structures. Acta Crystallogr D Biol Crystallogr. 1996 Jul 1;52(Pt 4):829–832. doi: 10.1107/S0907444996001783. [DOI] [PubMed] [Google Scholar]
  28. Kulkosky J., Jones K. S., Katz R. A., Mack J. P., Skalka A. M. Residues critical for retroviral integrative recombination in a region that is highly conserved among retroviral/retrotransposon integrases and bacterial insertion sequence transposases. Mol Cell Biol. 1992 May;12(5):2331–2338. doi: 10.1128/mcb.12.5.2331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lampe D. J., Churchill M. E., Robertson H. M. A purified mariner transposase is sufficient to mediate transposition in vitro. EMBO J. 1996 Oct 1;15(19):5470–5479. [PMC free article] [PubMed] [Google Scholar]
  30. Lamzin V. S., Wilson K. S. Automated refinement of protein models. Acta Crystallogr D Biol Crystallogr. 1993 Jan 1;49(Pt 1):129–147. doi: 10.1107/S0907444992008886. [DOI] [PubMed] [Google Scholar]
  31. Lavery R., Sklenar H. Defining the structure of irregular nucleic acids: conventions and principles. J Biomol Struct Dyn. 1989 Feb;6(4):655–667. doi: 10.1080/07391102.1989.10507728. [DOI] [PubMed] [Google Scholar]
  32. Morris A. L., MacArthur M. W., Hutchinson E. G., Thornton J. M. Stereochemical quality of protein structure coordinates. Proteins. 1992 Apr;12(4):345–364. doi: 10.1002/prot.340120407. [DOI] [PubMed] [Google Scholar]
  33. Murphy J. E., Goff S. P. A mutation at one end of Moloney murine leukemia virus DNA blocks cleavage of both ends by the viral integrase in vivo. J Virol. 1992 Aug;66(8):5092–5095. doi: 10.1128/jvi.66.8.5092-5095.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Murshudov G. N., Vagin A. A., Dodson E. J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr D Biol Crystallogr. 1997 May 1;53(Pt 3):240–255. doi: 10.1107/S0907444996012255. [DOI] [PubMed] [Google Scholar]
  35. 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]
  36. Ogata K., Morikawa S., Nakamura H., Hojo H., Yoshimura S., Zhang R., Aimoto S., Ametani Y., Hirata Z., Sarai A. Comparison of the free and DNA-complexed forms of the DNA-binding domain from c-Myb. Nat Struct Biol. 1995 Apr;2(4):309–320. doi: 10.1038/nsb0495-309. [DOI] [PubMed] [Google Scholar]
  37. Plasterk R. H. The Tc1/mariner transposon family. Curr Top Microbiol Immunol. 1996;204:125–143. doi: 10.1007/978-3-642-79795-8_6. [DOI] [PubMed] [Google Scholar]
  38. Rice P., Mizuuchi K. Structure of the bacteriophage Mu transposase core: a common structural motif for DNA transposition and retroviral integration. Cell. 1995 Jul 28;82(2):209–220. doi: 10.1016/0092-8674(95)90308-9. [DOI] [PubMed] [Google Scholar]
  39. Rost B., Sander C. Prediction of protein secondary structure at better than 70% accuracy. J Mol Biol. 1993 Jul 20;232(2):584–599. doi: 10.1006/jmbi.1993.1413. [DOI] [PubMed] [Google Scholar]
  40. Savilahti H., Mizuuchi K. Mu transpositional recombination: donor DNA cleavage and strand transfer in trans by the Mu transposase. Cell. 1996 Apr 19;85(2):271–280. doi: 10.1016/s0092-8674(00)81103-4. [DOI] [PubMed] [Google Scholar]
  41. Stofer E., Lavery R. Measuring the geometry of DNA grooves. Biopolymers. 1994 Mar;34(3):337–346. doi: 10.1002/bip.360340305. [DOI] [PubMed] [Google Scholar]
  42. Strauss J. K., Maher L. J., 3rd DNA bending by asymmetric phosphate neutralization. Science. 1994 Dec 16;266(5192):1829–1834. doi: 10.1126/science.7997878. [DOI] [PubMed] [Google Scholar]
  43. Suzuki M. Common features in DNA recognition helices of eukaryotic transcription factors. EMBO J. 1993 Aug;12(8):3221–3226. doi: 10.1002/j.1460-2075.1993.tb05991.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Suzuki M., Gerstein M. Binding geometry of alpha-helices that recognize DNA. Proteins. 1995 Dec;23(4):525–535. doi: 10.1002/prot.340230407. [DOI] [PubMed] [Google Scholar]
  45. Tullius T. Homeodomains: together again for the first time. Structure. 1995 Nov 15;3(11):1143–1145. doi: 10.1016/s0969-2126(01)00250-7. [DOI] [PubMed] [Google Scholar]
  46. Vos J. C., De Baere I., Plasterk R. H. Transposase is the only nematode protein required for in vitro transposition of Tc1. Genes Dev. 1996 Mar 15;10(6):755–761. doi: 10.1101/gad.10.6.755. [DOI] [PubMed] [Google Scholar]
  47. Vos J. C., Plasterk R. H. Tc1 transposase of Caenorhabditis elegans is an endonuclease with a bipartite DNA binding domain. EMBO J. 1994 Dec 15;13(24):6125–6132. doi: 10.1002/j.1460-2075.1994.tb06959.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Vos J. C., van Luenen H. G., Plasterk R. H. Characterization of the Caenorhabditis elegans Tc1 transposase in vivo and in vitro. Genes Dev. 1993 Jul;7(7A):1244–1253. doi: 10.1101/gad.7.7a.1244. [DOI] [PubMed] [Google Scholar]
  49. Wilson D. S., Guenther B., Desplan C., Kuriyan J. High resolution crystal structure of a paired (Pax) class cooperative homeodomain dimer on DNA. Cell. 1995 Sep 8;82(5):709–719. doi: 10.1016/0092-8674(95)90468-9. [DOI] [PubMed] [Google Scholar]
  50. Wintjens R., Rooman M. Structural classification of HTH DNA-binding domains and protein-DNA interaction modes. J Mol Biol. 1996 Sep 20;262(2):294–313. doi: 10.1006/jmbi.1996.0514. [DOI] [PubMed] [Google Scholar]
  51. Xu W., Rould M. A., Jun S., Desplan C., Pabo C. O. Crystal structure of a paired domain-DNA complex at 2.5 A resolution reveals structural basis for Pax developmental mutations. Cell. 1995 Feb 24;80(4):639–650. doi: 10.1016/0092-8674(95)90518-9. [DOI] [PubMed] [Google Scholar]
  52. Yang J. Y., Jayaram M., Harshey R. M. Positional information within the Mu transposase tetramer: catalytic contributions of individual monomers. Cell. 1996 May 3;85(3):447–455. doi: 10.1016/s0092-8674(00)81122-8. [DOI] [PubMed] [Google Scholar]
  53. Yang W., Hendrickson W. A., Crouch R. J., Satow Y. Structure of ribonuclease H phased at 2 A resolution by MAD analysis of the selenomethionyl protein. Science. 1990 Sep 21;249(4975):1398–1405. doi: 10.1126/science.2169648. [DOI] [PubMed] [Google Scholar]
  54. Yang W., Steitz T. A. Recombining the structures of HIV integrase, RuvC and RNase H. Structure. 1995 Feb 15;3(2):131–134. doi: 10.1016/s0969-2126(01)00142-3. [DOI] [PubMed] [Google Scholar]
  55. van Luenen H. G., Colloms S. D., Plasterk R. H. Mobilization of quiet, endogenous Tc3 transposons of Caenorhabditis elegans by forced expression of Tc3 transposase. EMBO J. 1993 Jun;12(6):2513–2520. doi: 10.1002/j.1460-2075.1993.tb05906.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. van Luenen H. G., Colloms S. D., Plasterk R. H. The mechanism of transposition of Tc3 in C. elegans. Cell. 1994 Oct 21;79(2):293–301. doi: 10.1016/0092-8674(94)90198-8. [DOI] [PubMed] [Google Scholar]

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