Structure of the VirB4 ATPase, alone and bound to the core complex of a type IV secretion system
10.1073/pnas.1201428109
Crossref journal-article
Proceedings of the National Academy of Sciences
Proceedings of the National Academy of Sciences (341)
Abstract

Type IV secretion (T4S) systems mediate the transfer of proteins and DNA across the cell envelope of bacteria. These systems play important roles in bacterial pathogenesis and in horizontal transfer of antibiotic resistance. The VirB4 ATPase of the T4S system is essential for both the assembly of the system and substrate transfer. In this article, we present the crystal structure of the C-terminal domain of Thermoanaerobacter pseudethanolicus VirB4. This structure is strikingly similar to that of another T4S ATPase, VirD4, a protein that shares only 12% sequence identity with VirB4. The VirB4 domain purifies as a monomer, but the full-length protein is observed in a monomer-dimer equilibrium, even in the presence of nucleotides and DNAs. We also report the negative stain electron microscopy structure of the core complex of the T4S system of the Escherichia coli pKM101 plasmid, with VirB4 bound. In this structure, VirB4 is also monomeric and bound through its N-terminal domain to the core’s VirB9 protein. Remarkably, VirB4 is observed bound to the side of the complex where it is ideally placed to play its known regulatory role in substrate transfer.

Bibliography

Walldén, K., Williams, R., Yan, J., Lian, P. W., Wang, L., Thalassinos, K., Orlova, E. V., & Waksman, G. (2012). Structure of the VirB4 ATPase, alone and bound to the core complex of a type IV secretion system. Proceedings of the National Academy of Sciences, 109(28), 11348–11353.

Authors 8
  1. Karin Walldén (first)
  2. Robert Williams (additional)
  3. Jun Yan (additional)
  4. Pei W. Lian (additional)
  5. Luchun Wang (additional)
  6. Konstantinos Thalassinos (additional)
  7. Elena V. Orlova (additional)
  8. Gabriel Waksman (additional)
References 45 Referenced 80
  1. E Grohmann, G Muth, M Espinosa, Conjugative plasmid transfer in gram-positive bacteria. Microbiol Mol Biol Rev 67, 277–301 (2003). (10.1128/MMBR.67.2.277-301.2003) / Microbiol Mol Biol Rev / Conjugative plasmid transfer in gram-positive bacteria by Grohmann E (2003)
  2. K Wallden, A Rivera-Calzada, G Waksman, Type IV secretion systems: Versatility and diversity in function. Cell Microbiol 12, 1203–1212 (2010). (10.1111/j.1462-5822.2010.01499.x) / Cell Microbiol / Type IV secretion systems: Versatility and diversity in function by Wallden K (2010)
  3. T Kwok, et al., Helicobacter exploits integrin for type IV secretion and kinase activation. Nature 449, 862–866 (2007). (10.1038/nature06187) / Nature / Helicobacter exploits integrin for type IV secretion and kinase activation by Kwok T (2007)
  4. Y Zhao, et al., Role of a type IV-like secretion system of Streptococcus suis 2 in the development of streptococcal toxic shock syndrome. J Infect Dis 204, 274–281 (2011). (10.1093/infdis/jir261) / J Infect Dis / Role of a type IV-like secretion system of Streptococcus suis 2 in the development of streptococcal toxic shock syndrome by Zhao Y (2011)
  5. PJ Christie, K Atmakuri, V Krishnamoorthy, S Jakubowski, E Cascales, Biogenesis, architecture, and function of bacterial type IV secretion systems. Annu Rev Microbiol 59, 451–485 (2005). (10.1146/annurev.micro.58.030603.123630) / Annu Rev Microbiol / Biogenesis, architecture, and function of bacterial type IV secretion systems by Christie PJ (2005)
  6. R Fronzes, et al., Structure of a type IV secretion system core complex. Science 323, 266–268 (2009). (10.1126/science.1166101) / Science / Structure of a type IV secretion system core complex by Fronzes R (2009)
  7. V Chandran, et al., Structure of the outer membrane complex of a type IV secretion system. Nature 462, 1011–1015 (2009). (10.1038/nature08588) / Nature / Structure of the outer membrane complex of a type IV secretion system by Chandran V (2009)
  8. R Eisenbrandt, et al., Conjugative pili of IncP plasmids, and the Ti plasmid T pilus are composed of cyclic subunits. J Biol Chem 274, 22548–22555 (1999). (10.1074/jbc.274.32.22548) / J Biol Chem / Conjugative pili of IncP plasmids, and the Ti plasmid T pilus are composed of cyclic subunits by Eisenbrandt R (1999)
  9. KA Aly, C Baron, The VirB5 protein localizes to the T-pilus tips in Agrobacterium tumefaciens. Microbiology 153, 3766–3775 (2007). (10.1099/mic.0.2007/010462-0) / Microbiology / The VirB5 protein localizes to the T-pilus tips in Agrobacterium tumefaciens by Aly KA (2007)
  10. R Fronzes, PJ Christie, G Waksman, The structural biology of type IV secretion systems. Nat Rev Microbiol 7, 703–714 (2009). (10.1038/nrmicro2218) / Nat Rev Microbiol / The structural biology of type IV secretion systems by Fronzes R (2009)
  11. HJ Yeo, SN Savvides, AB Herr, E Lanka, G Waksman, Crystal structure of the hexameric traffic ATPase of the Helicobacter pylori type IV secretion system. Mol Cell 6, 1461–1472 (2000). (10.1016/S1097-2765(00)00142-8) / Mol Cell / Crystal structure of the hexameric traffic ATPase of the Helicobacter pylori type IV secretion system by Yeo HJ (2000)
  12. FX Gomis-Rüth, et al., The bacterial conjugation protein TrwB resembles ring helicases and F1-ATPase. Nature 409, 637–641 (2001). (10.1038/35054586) / Nature / The bacterial conjugation protein TrwB resembles ring helicases and F1-ATPase by Gomis-Rüth FX (2001)
  13. S Hare, R Bayliss, C Baron, G Waksman, A large domain swap in the VirB11 ATPase of Brucella suis leaves the hexameric assembly intact. J Mol Biol 360, 56–66 (2006). (10.1016/j.jmb.2006.04.060) / J Mol Biol / A large domain swap in the VirB11 ATPase of Brucella suis leaves the hexameric assembly intact by Hare S (2006)
  14. KJ Fullner, JC Lara, EW Nester, Pilus assembly by Agrobacterium T-DNA transfer genes. Science 273, 1107–1109 (1996). (10.1126/science.273.5278.1107) / Science / Pilus assembly by Agrobacterium T-DNA transfer genes by Fullner KJ (1996)
  15. M Watarai, S Makino, T Shirahata, An essential virulence protein of Brucella abortus, VirB4, requires an intact nucleoside-triphosphate-binding domain. Microbiology 148, 1439–1446 (2002). (10.1099/00221287-148-5-1439) / Microbiology / An essential virulence protein of Brucella abortus, VirB4, requires an intact nucleoside-triphosphate-binding domain by Watarai M (2002)
  16. BR Berger, PJ Christie, The Agrobacterium tumefaciens virB4 gene product is an essential virulence protein requiring an intact nucleoside triphosphate-binding domain. J Bacteriol 175, 1723–1734 (1993). (10.1128/jb.175.6.1723-1734.1993) / J Bacteriol / The Agrobacterium tumefaciens virB4 gene product is an essential virulence protein requiring an intact nucleoside triphosphate-binding domain by Berger BR (1993)
  17. KJ Fullner, KM Stephens, EW Nester, An essential virulence protein of Agrobacterium tumefaciens, VirB4, requires an intact mononucleotide binding domain to function in transfer of T-DNA. Mol Gen Genet 245, 704–715 (1994). (10.1007/BF00297277) / Mol Gen Genet / An essential virulence protein of Agrobacterium tumefaciens, VirB4, requires an intact mononucleotide binding domain to function in transfer of T-DNA by Fullner KJ (1994)
  18. R Middleton, K Sjölander, N Krishnamurthy, J Foley, P Zambryski, Predicted hexameric structure of the Agrobacterium VirB4 C terminus suggests VirB4 acts as a docking site during type IV secretion. Proc Natl Acad Sci USA 102, 1685–1690 (2005). (10.1073/pnas.0409399102) / Proc Natl Acad Sci USA / Predicted hexameric structure of the Agrobacterium VirB4 C terminus suggests VirB4 acts as a docking site during type IV secretion by Middleton R (2005)
  19. Q Yuan, et al., Identification of the VirB4-VirB8-VirB5-VirB2 pilus assembly sequence of type IV secretion systems. J Biol Chem 280, 26349–26359 (2005). (10.1074/jbc.M502347200) / J Biol Chem / Identification of the VirB4-VirB8-VirB5-VirB2 pilus assembly sequence of type IV secretion systems by Yuan Q (2005)
  20. I Arechaga, et al., ATPase activity and oligomeric state of TrwK, the VirB4 homologue of the plasmid R388 type IV secretion system. J Bacteriol 190, 5472–5479 (2008). (10.1128/JB.00321-08) / J Bacteriol / ATPase activity and oligomeric state of TrwK, the VirB4 homologue of the plasmid R388 type IV secretion system by Arechaga I (2008)
  21. C Rabel, AM Grahn, R Lurz, E Lanka, The VirB4 family of proposed traffic nucleoside triphosphatases: Common motifs in plasmid RP4 TrbE are essential for conjugation and phage adsorption. J Bacteriol 185, 1045–1058 (2003). (10.1128/JB.185.3.1045-1058.2003) / J Bacteriol / The VirB4 family of proposed traffic nucleoside triphosphatases: Common motifs in plasmid RP4 TrbE are essential for conjugation and phage adsorption by Rabel C (2003)
  22. TA Dang, XR Zhou, B Graf, PJ Christie, Dimerization of the Agrobacterium tumefaciens VirB4 ATPase and the effect of ATP-binding cassette mutations on the assembly and function of the T-DNA transporter. Mol Microbiol 32, 1239–1253 (1999). (10.1046/j.1365-2958.1999.01436.x) / Mol Microbiol / Dimerization of the Agrobacterium tumefaciens VirB4 ATPase and the effect of ATP-binding cassette mutations on the assembly and function of the T-DNA transporter by Dang TA (1999)
  23. E Durand, C Oomen, G Waksman, Biochemical dissection of the ATPase TraB, the VirB4 homologue of the Escherichia coli pKM101 conjugation machinery. J Bacteriol 192, 2315–2323 (2010). (10.1128/JB.01384-09) / J Bacteriol / Biochemical dissection of the ATPase TraB, the VirB4 homologue of the Escherichia coli pKM101 conjugation machinery by Durand E (2010)
  24. E Durand, G Waksman, V Receveur-Brechot, Structural insights into the membrane-extracted dimeric form of the ATPase TraB from the Escherichia coli pKM101 conjugation system. BMC Struct Biol 11, 4 (2011). (10.1186/1472-6807-11-4) / BMC Struct Biol / Structural insights into the membrane-extracted dimeric form of the ATPase TraB from the Escherichia coli pKM101 conjugation system by Durand E (2011)
  25. O Draper, R Middleton, M Doucleff, PC Zambryski, Topology of the VirB4 C terminus in the Agrobacterium tumefaciens VirB/D4 type IV secretion system. J Biol Chem 281, 37628–37635 (2006). (10.1074/jbc.M606403200) / J Biol Chem / Topology of the VirB4 C terminus in the Agrobacterium tumefaciens VirB/D4 type IV secretion system by Draper O (2006)
  26. MY Abajy, et al., A type IV-secretion-like system is required for conjugative DNA transport of broad-host-range plasmid pIP501 in gram-positive bacteria. J Bacteriol 189, 2487–2496 (2007). (10.1128/JB.01491-06) / J Bacteriol / A type IV-secretion-like system is required for conjugative DNA transport of broad-host-range plasmid pIP501 in gram-positive bacteria by Abajy MY (2007)
  27. JA Caryl, AJ O’Neill, Complete nucleotide sequence of pGO1, the prototype conjugative plasmid from the Staphylococci. Plasmid 62, 35–38 (2009). (10.1016/j.plasmid.2009.03.001) / Plasmid / Complete nucleotide sequence of pGO1, the prototype conjugative plasmid from the Staphylococci by Caryl JA (2009)
  28. TD Lawley, WA Klimke, MJ Gubbins, LS Frost, F factor conjugation is a true type IV secretion system. FEMS Microbiol Lett 224, 1–15 (2003). (10.1016/S0378-1097(03)00430-0) / FEMS Microbiol Lett / F factor conjugation is a true type IV secretion system by Lawley TD (2003)
  29. KA Schandel, MM Muller, RE Webster, Localization of TraC, a protein involved in assembly of the F conjugative pilus. J Bacteriol 174, 3800–3806 (1992). (10.1128/jb.174.11.3800-3806.1992) / J Bacteriol / Localization of TraC, a protein involved in assembly of the F conjugative pilus by Schandel KA (1992)
  30. RL Woodbury, SJ Hardy, LL Randall, Complex behavior in solution of homodimeric SecA. Protein Sci 11, 875–882 (2002). (10.1110/ps.4090102) / Protein Sci / Complex behavior in solution of homodimeric SecA by Woodbury RL (2002)
  31. S Korolev, J Hsieh, GH Gauss, TM Lohman, G Waksman, Major domain swiveling revealed by the crystal structures of complexes of E. coli Rep helicase bound to single-stranded DNA and ADP. Cell 90, 635–647 (1997). (10.1016/S0092-8674(00)80525-5) / Cell / Major domain swiveling revealed by the crystal structures of complexes of E. coli Rep helicase bound to single-stranded DNA and ADP by Korolev S (1997)
  32. K Kazetani, T Minamino, T Miyata, T Kato, K Namba, ATP-induced FliI hexamerization facilitates bacterial flagellar protein export. Biochem Biophys Res Commun 388, 323–327 (2009). (10.1016/j.bbrc.2009.08.004) / Biochem Biophys Res Commun / ATP-induced FliI hexamerization facilitates bacterial flagellar protein export by Kazetani K (2009)
  33. PA Tucker, L Sallai, The AAA+ superfamily—A myriad of motions. Curr Opin Struct Biol 17, 641–652 (2007). (10.1016/j.sbi.2007.09.012) / Curr Opin Struct Biol / The AAA+ superfamily—A myriad of motions by Tucker PA (2007)
  34. HD de Paz, et al., Functional dissection of the conjugative coupling protein TrwB. J Bacteriol 192, 2655–2669 (2010). (10.1128/JB.01692-09) / J Bacteriol / Functional dissection of the conjugative coupling protein TrwB by de Paz HD (2010)
  35. EF Pettersen, et al., UCSF Chimera—A visualization system for exploratory research and analysis. J Comput Chem 25, 1605–1612 (2004). (10.1002/jcc.20084) / J Comput Chem / UCSF Chimera—A visualization system for exploratory research and analysis by Pettersen EF (2004)
  36. J Zimmer, Y Nam, TA Rapoport, Structure of a complex of the ATPase SecA and the protein-translocation channel. Nature 455, 936–943 (2008). (10.1038/nature07335) / Nature / Structure of a complex of the ATPase SecA and the protein-translocation channel by Zimmer J (2008)
  37. AJ Heck, Native mass spectrometry: A bridge between interactomics and structural biology. Nat Methods 5, 927–933 (2008). (10.1038/nmeth.1265) / Nat Methods / Native mass spectrometry: A bridge between interactomics and structural biology by Heck AJ (2008)
  38. HR Bourne, G proteins. The arginine finger strikes again. Nature 389, 673–674 (1997). (10.1038/39470) / Nature / G proteins. The arginine finger strikes again by Bourne HR (1997)
  39. E Cascales, PJ Christie, Definition of a bacterial type IV secretion pathway for a DNA substrate. Science 304, 1170–1173 (2004). (10.1126/science.1095211) / Science / Definition of a bacterial type IV secretion pathway for a DNA substrate by Cascales E (2004)
  40. P Mossey, A Hudacek, A Das, Agrobacterium tumefaciens type IV secretion protein VirB3 is an inner membrane protein and requires VirB4, VirB7, and VirB8 for stabilization. J Bacteriol 192, 2830–2838 (2010). (10.1128/JB.01331-09) / J Bacteriol / Agrobacterium tumefaciens type IV secretion protein VirB3 is an inner membrane protein and requires VirB4, VirB7, and VirB8 for stabilization by Mossey P (2010)
  41. L Terradot, et al., Biochemical characterization of protein complexes from the Helicobacter pylori protein interaction map: strategies for complex formation and evidence for novel interactions within type IV secretion systems. Mol Cell Proteomics 3, 809–819 (2004). (10.1074/mcp.M400048-MCP200) / Mol Cell Proteomics / Biochemical characterization of protein complexes from the Helicobacter pylori protein interaction map: strategies for complex formation and evidence for novel interactions within type IV secretion systems by Terradot L (2004)
  42. E Cascales, PJ Christie, Agrobacterium VirB10, an ATP energy sensor required for type IV secretion. Proc Natl Acad Sci USA 101, 17228–17233 (2004). (10.1073/pnas.0405843101) / Proc Natl Acad Sci USA / Agrobacterium VirB10, an ATP energy sensor required for type IV secretion by Cascales E (2004)
  43. PD Adams, et al., PHENIX: A comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr D Biol Crystallogr 66, 213–221 (2010). (10.1107/S0907444909052925) / Acta Crystallogr D Biol Crystallogr / PHENIX: A comprehensive Python-based system for macromolecular structure solution by Adams PD (2010)
  44. SD Pringle, et al., An investigation of the mobility separation of some peptide and protein ions using a new hybrid quadrupole/travelling wave IMS/oa-ToF instrument. Int J Mass Spectrom 261, 1–12 (2007). (10.1016/j.ijms.2006.07.021) / Int J Mass Spectrom / An investigation of the mobility separation of some peptide and protein ions using a new hybrid quadrupole/travelling wave IMS/oa-ToF instrument by Pringle SD (2007)
  45. M van Heel, et al., Single-particle electron cryo-microscopy: Towards atomic resolution. Q Rev Biophys 33, 307–369 (2000). (10.1017/S0033583500003644) / Q Rev Biophys / Single-particle electron cryo-microscopy: Towards atomic resolution by van Heel M (2000)
Dates
Type When
Created 13 years, 1 month ago (June 28, 2012, 8:19 p.m.)
Deposited 3 years, 2 months ago (June 7, 2022, 4:45 a.m.)
Indexed 1 year, 1 month ago (July 17, 2024, 4:28 a.m.)
Issued 13 years, 1 month ago (June 27, 2012)
Published 13 years, 1 month ago (June 27, 2012)
Published Online 13 years, 1 month ago (June 27, 2012)
Published Print 13 years, 1 month ago (July 10, 2012)
Funders 0

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@article{Walld_n_2012, title={Structure of the VirB4 ATPase, alone and bound to the core complex of a type IV secretion system}, volume={109}, ISSN={1091-6490}, url={http://dx.doi.org/10.1073/pnas.1201428109}, DOI={10.1073/pnas.1201428109}, number={28}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Walldén, Karin and Williams, Robert and Yan, Jun and Lian, Pei W. and Wang, Luchun and Thalassinos, Konstantinos and Orlova, Elena V. and Waksman, Gabriel}, year={2012}, month=jun, pages={11348–11353} }