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Differential processing of quorum sensing signals through phosphotransfer: structural insights from molecular dynamics simulations

  • Devlina Chakravarty
  • Pinak ChakrabartiEmail author
  • Mousumi BanerjeeEmail author
Original Article
  • 6 Downloads

Abstract

The auto-inducer-mediated virulence gene expression and biofilm formation in Vibrio sp. uses a highly evolved two-component phosphotransfer system, involving a histidine sensor kinase (LuxQ), an Hpt domain protein (LuxU) and a universal response regulator (LuxO), to process the signal. At low and high cell density, the phosphotransfer reaction occurs differently leading to the activation or deactivation, respectively, of the global repressor which in turn regulates the virulence. Here the molecular details of signal processing and signal decay have been studied using structural modelling and molecular dynamics simulation of LuxQ, LuxU and LuxO individual proteins and protein–protein complexes with and without the phosphate group. The stability, conformational flexibility and structural changes associated with phosphotransfer of the individual protein and the protein–protein complexes are compared. The root mean square deviations and the root mean square fluctuations of the phosphorylated and unphosphorylated proteins showed significant differences in these two processes. The principal component analysis points out the remarkable differences in the essential motions of the systems, which depend not only on the phosphorylated complex but also on the key phosphorylation of the individual protein component. This observation is also highlighted by the dynamic cross-correlation matrix (DCCM) analysis where concerted motions are found to differ depending on the state of phosphorylation. Evaluation of the equilibrated structures and their free energy reveals that the reverse transfer of phosphate during signal decay is energetically less favourable.

Keywords

Vibrio harveyi Quorum sensing Auto-inducer Signalling pathway proteins Molecular dynamics simulation 

Abbreviations

MD

Molecular dynamics

PDB

Protein Data Bank

NMR

Nuclear magnetic resonance

GBSA

Generalized Born solvent accessible surface area

RMSD

Root mean square deviation

RMSF

Root mean square fluctuation

DCCM

Dynamic cross-correlation map

PCA

Principal component analysis

Notes

Acknowledgements

Our sincere thanks to Drs. Soumalee Basu and Shaon Roy Choudhuri (Maulana Abul Kalam Azad University of Technology) for their helpful discussion. MB thanks UGC, India, for D. S Kothari post-doctoral fellowship and Dr. Raju Mukherjee for a critical reading of the manuscript.

Compliance with ethical standards

Conflict of interest

The authors have no conflict of interest.

Supplementary material

42485_2019_10_MOESM1_ESM.docx (3.5 mb)
Supplementary material 1 (DOCX 3552 kb)
42485_2019_10_MOESM2_ESM.mpg (5.6 mb)
Supplementary material 2 (MPG 5698 kb) SM1. Dynamics of unphosphorylated LuxUQ. LuxU is shown in green and LuxQ in blue cartoons, active site residues His58 in blue and Asp107 in red sticks
42485_2019_10_MOESM3_ESM.mpg (6 mb)
Supplementary material 3 (MPG 6170 kb) SM2. Dynamics of LuxUQ with phosphorylated Asp107
42485_2019_10_MOESM4_ESM.mpg (5.2 mb)
Supplementary material 4 (MPG 5306 kb) SM3. Dynamics of LuxUQ with phosphorylated His58
42485_2019_10_MOESM5_ESM.jpg (55 kb)
Supplementary material 5 (JPEG 54 kb) SM4. The motion captured by PC1 in unphosphorylated LuxUQ. The protein is shown as ribbon (LuxQ in blue). The helices in LuxU are demarcated using different colors, H1 (Asn7-Leu44, red), H2 (Gly46-Phe67, magenta), H3 (Ala69 to Leu88, yellow) and H4 (Glu97 to Ser111, blue). His58 in helix H2 is represented by a sphere
42485_2019_10_MOESM6_ESM.jpg (55 kb)
Supplementary material 6 (JPEG 54 kb) SM5. The motion captured by PC1 in LuxUQ with phosphorylated Asp107 (LuxQ in blue, and LuxU in green)
42485_2019_10_MOESM7_ESM.jpg (65 kb)
Supplementary material 7 (JPEG 64 kb) SM6. The motion captured by PC1 in LuxUQ with phosphorylated His58
42485_2019_10_MOESM8_ESM.mpg (5.1 mb)
Supplementary material 8 (MPG 5222 kb) SM7. Dynamics of unphosphorylated LuxUO. LuxU is shown in green and LuxO in orange cartoons, active site residues His58 in blue and Asp66 in red sticks
42485_2019_10_MOESM9_ESM.mpg (5.3 mb)
Supplementary material 9 (MPG 5435 kb) SM8. Dynamics of LuxUO with phosphorylated His58
42485_2019_10_MOESM10_ESM.mpg (4.2 mb)
Supplementary material 10 (MPG 4337 kb) SM9. Dynamics of LuxUO with phosphorylated Asp66
42485_2019_10_MOESM11_ESM.jpg (53 kb)
Supplementary material 11 (JPEG 53 kb) SM10. The motion captured by PC1 in unphosphorylated LuxUO. The protein backbone (Cα trace) is shown as ribbon (LuxU in green and LuxO in orange)
42485_2019_10_MOESM12_ESM.jpg (54 kb)
Supplementary material 12 (JPEG 53 kb) SM11. The motion captured by PC1 in LuxUO with phosphorylated His58
42485_2019_10_MOESM13_ESM.jpg (57 kb)
Supplementary material 13 (JPEG 57 kb) SM12. The motion captured by PC1 in LuxUO with phosphorylated Asp66

References

  1. Aguilar PS, Hernandez-Arriaga AM, Cybulski LE, Erazo AC, de Mendoza D (2001) Molecular basis of thermosensing: a two-component signal transduction thermometer in bascillussubtilis. EMBO J 20(7):1681–1691CrossRefGoogle Scholar
  2. Albanesi D, Martín M, Trajtenberg F, Mansilla MC, Haouz A, Alzari PM, de Mendoza D, Buschiazzo A (2009) Structural plasticity and catalysis regulation of thermosensor histidine kinase. Proc Natl Acad Sci USA 106(38):16185–16190CrossRefGoogle Scholar
  3. Ashkenazy H, Erez E, Martz E, Pupko T, Ben-Tal N (2010) ConSurf 2010: calculating evolutionary conservation in sequence and structure of proteins and nucleic acids. Acids Res, Nucl.  https://doi.org/10.1093/nar/gkq399 Google Scholar
  4. Ashkenazy H, Abadi S, Martz E, Chay O, Mayrose I, Pupko T, Ben-Tal N (2016) ConSurf 2016: an improved methodology to estimate and visualize evolutionary conservation in macromolecules. Nucleic Acids Res 44(W1):W344–W350CrossRefGoogle Scholar
  5. Bassler BL, Losick R (2006) Bacterially speaking. Cell 125(2):237–246CrossRefGoogle Scholar
  6. Bhate MP, Molnar KS, Goulian M, DeGrado WF (2015) Signal transduction in histidine kinases: insights from new structures. Structure 23(6):981–994CrossRefGoogle Scholar
  7. Boyaci H, Shah T, Hurley A, Kokona B, Li Z, Ventocilla C, Jeffrey PD, Semmelhack MF, Fairman R, Bassler BL, Hughson FM (2016) Structure, regulation and inhibition of the quorum sensing signal integrator LuxO. PLoS Biol 14:e1002464CrossRefGoogle Scholar
  8. Cámara M, Williams P, Hardman A (2002) Controlling infection by tuning in and tuning down the volume of bacterial small talk. Lancet Infect Dis 2(11):667–676CrossRefGoogle Scholar
  9. Capra EJ, Laub MT (2012) Evolution of two-component signal transduction systems. Annu Rev Microbiol 66:325–347CrossRefGoogle Scholar
  10. Case et al (2005) The AMBER biomolecular simulation program. AMBER 10.0. J Comput Chem 26(16):1668–1688CrossRefGoogle Scholar
  11. Celniker G, Nimrod G, Ashkenazy H, Glaser F, Martz E, Mayrose I, Pupko T, Ben-Tal N (2013) ConSurf: using evolutionary data to raise testable hypotheses about protein function. Isr J Chem 53(3–4):199–206CrossRefGoogle Scholar
  12. Cheung J, Hendrickson WA (2009) Structural analysis and ligand simulation of the histidine kinase NarX. Structure 17(2):190–201CrossRefGoogle Scholar
  13. Faloon P, Weiner WS, Matharu DS, Neuenswander B, Porubsky P, Youngsaye W, Bennion M, Ng WL, Hurley A, Mosher CM, Johnston S, Dandapani S, Schoenen FJ, Aubé J, Munoz B, Palmer M, Bassler BL, Schreiber SL (2010–2013a) Probe reports from the NIH Molecular Libraries Program. National Center for Biotechnology Information (US), Bethesda (discovery of ML370, an inhibitor of Vibrio cholerae quorum sensing acting via the LuxO response regulator) Google Scholar
  14. Faloon P, Jewett I, Youngsaye W, Bennion M, Ng WL, Hurley A, Lewis, TA, Edwankar RV, Le H, Mosher CM, Johnston S, Dandapani S, Munoz B, Palmer M, Bassler BL, Schreiber SL (2010–2013b) Probe reports from the NIH Molecular Libraries Program. National Center for Biotechnology Information (US), Bethesda (discovery of ML366, an inhibitor of Vibrio cholerae quorum sensing acting via the LuxO response regulator) Google Scholar
  15. Feng L, Rutherford ST, Papenfort K, Bagert JD, van Kessel JC, Tirrell DA, Wingreen NS, Bassler BL (2015) A qrr noncoding RNA deploys four different regulatory mechanisms to optimize quorum-sensing dynamics. Cell 160(1–2):228–240CrossRefGoogle Scholar
  16. Freeman JA, Bassler BL (1999) A genetic analysis of the function of LuxO, a two –component response regulator involved in quorum sensing in Vibrio harveyi. Mol Microbiol 31(2):665–677CrossRefGoogle Scholar
  17. Glaser F, Pupko T, Paz I, Bell RE, Bechor-Shental D, Martz E, Ben-Tal N (2003) ConSurf: identification of functional regions in proteins by surface-mapping of phylogenetic information. Bioinformatics 19(1):163–164CrossRefGoogle Scholar
  18. Grant BJ, Rodrigues APC, ElSawy KM, McCammon JA, Caves LSD (2006) Bio3d: an R package for the comparative analysis of protein structures. Bioinformatics 22(21):2695–2696CrossRefGoogle Scholar
  19. Groisman Eduardo A (2001) The pleiotropic two-component regulatory system PhoP–PhoQ. J Bacteriol 183(6):1835–1842CrossRefGoogle Scholar
  20. Henke JM, Bassler BL (2004a) Three parallel quorum-sensing systems regulate gene expression in Vibrio harveyi. J Bacteriol 186(20):6902–6914CrossRefGoogle Scholar
  21. Henke JM, Bassler BL (2004b) Bacterial social engagement. Trends Cell Biol 14(11):648–656CrossRefGoogle Scholar
  22. Humphrey W, Dalke A, Schulten K (1996) VMD: visual molecular dynamics. J Mol Graph 14:33–38CrossRefGoogle Scholar
  23. Landau M, Mayrose I, Rosenberg Y, Glaser F, Martz E, Pupko T, Ben-Tal N (2005) ConSurf 2005: the projection of evolutionary conservation scores of residues on protein structures. Nucleic Acids Res 33(suppl_2):W299–W302CrossRefGoogle Scholar
  24. Laub MT, Goulian M (2007) Specificity in two-component signal transduction pathways. Annu Rev Genet 41:121–145CrossRefGoogle Scholar
  25. Lauria A, Ippolito M, Almerico AM (2009) Principal component analysis on molecular descriptors as an alternative point of view in the search of new Hsp90 inhibitors. Comput Biol Chem 33:386–390CrossRefGoogle Scholar
  26. Lobanov MY, Ogatyrev NS, Galzitskaya OV (2008) Radius of gyration as an indicator of protein structure compactness. Mol Biol 42(4):623–628CrossRefGoogle Scholar
  27. Miller MB, Bassler BL (2001) Quorum sensing in bacteria. Annu Rev Microbiol 55:165–199CrossRefGoogle Scholar
  28. Neiditch MB, Federle MJ, Miller ST, Bassler BL, Hughson FM (2005) Regulation of LuxPQ receptor activity by the quorum-sensing signal autoinducer-2. Mol Cell 18(5):507–518CrossRefGoogle Scholar
  29. Neiditch MB, Federle MJ, Pompeani AJ, Kelly RC, Swem DL, Jeffrey PD, Bassler BL, Hughson FM (2006) Ligand-induced asymmetry in histidine sensor kinase complex regulates quorum sensing. Cell 126(6):1095–1108CrossRefGoogle Scholar
  30. Ng WL, Bassler BL (2009) Bacterial quorum-sensing network architecture. Annu Rev Genet 43:197–222CrossRefGoogle Scholar
  31. Onufriev A, Case DA, Bashford D (2002) Effective born radii in the generalized born approximation: the importance of being perfect. J Comput Chem 23(14):1297–1304CrossRefGoogle Scholar
  32. Papenfort K, Bassler BL (2016) Quorum sensing signal-response systems in Gram-negative bacteria. Nat Rev Microbiol 14(9):576–588CrossRefGoogle Scholar
  33. Papenfort K, Förstner KU, Cong JP, Sharma CM, Bassler BL (2015) Differential RNA-seq of Vibrio cholera identifies the VqmR small RNA as a regulator of biofilm formation. Proc Natl Acad Sci USA 112(7):E766–E775CrossRefGoogle Scholar
  34. Patskovsky Y, Ramagopal UA, Fong R, Freeman J, Iizuka M, Groshong C, Smith D, Wasserman SR, Sauder JM, Burley SK, Almo SC (2008) (to be published) Google Scholar
  35. Rutherford ST, Bassler BL (2012) Bacterial quorum sensing: its role in virulence and possibilities for its control. Cold Spring Harb Perspect Med 2(11):a012427CrossRefGoogle Scholar
  36. Rutherford ST, van Kessel JC, Shao Y, Bassler BL (2011) AphA and LuxR/HapR reciprocally control quorum sensing in vibrios. Ligand induced asymmetry in histidine sensor kinase complex regulates quorum sensing. Genes Dev 25(4):397–408CrossRefGoogle Scholar
  37. Salazar ME, Laub MT (2015) Temporal and evolutionary dynamics of two-component signalling pathways. Curr Opin Microbiol 24:7–14CrossRefGoogle Scholar
  38. Shao Y, Feng L, Rutherford ST, Papenfort K, Bassler BL (2013) Functional determinants of the quorum-sensing non-coding RNAs and their roles in target regulation. EMBO J 32(15):2158–2171CrossRefGoogle Scholar
  39. Stewart RC (2010) Protein histidine kinases: assembly of active sites and their regulation in signalling pathways. Curr Opin Microbiol 13(2):133–141CrossRefGoogle Scholar
  40. Stock AM, Robinson VL, Goudreau PN (2000) Two component signal transduction. Annu Rev Biochem 69:183–215CrossRefGoogle Scholar
  41. Svenningsen SL, Waters CM, Bassler BL (2008) A negative feedback loop involving small RNAs accelerates Vibrio cholera’s transition out of quorum sensing mode. Genes Dev 22(2):226–238CrossRefGoogle Scholar
  42. Swem LR, Swem DL, Wingreen NS, Bassler BL (2008) Deducing receptor signalling parameters from in vivo analysis: LuxN/A-1 quorum sensing in Vibrio harveyi. Cell 134(3):461–473CrossRefGoogle Scholar
  43. The PyMOL Molecular Graphics System, Version 1.8 (2019) Schrödinger, LLCGoogle Scholar
  44. Torre JG, Hueratas ML, Carrasco B (2000) Calculation of hydrodynamic properties of globular protein from their atomic level structure. Biophys J 78:719–730CrossRefGoogle Scholar
  45. Tovchigrechko A, Vakser IA (2006) GRAMM-X public web server for protein–protein docking. Nucleic Acids Res 34(Web Server issue):W310–W314CrossRefGoogle Scholar
  46. Tu KC, Bassler BL (2007) Multiple small RNAs act additively to integrate sensory information and control quorum sensing in Vibrio harveyi. Genes Dev 21(2):221–233CrossRefGoogle Scholar
  47. Tu KC, Waters CM, Svenningsen SL, Bassler BL (2008) A small RNA-mediated negative feedback loop controls quorum-sensing dynamics in Vibrio harveyi. Mol Microbiol 70(4):896–907Google Scholar
  48. Ulrich LE, Zulin IB (2010) The MisT2 database: a comprehensive genomics resource on microbial signal transduction. Nucleic Acid Res 38:D401–D407CrossRefGoogle Scholar
  49. Ulrich DL, Kojetin D, Bassler BL, Cavanagh J, Loria JP (2005) Solution structure and dynamics of LuxU from Vibrio harveyi, a phosphotransferase protein involved in bacterial quorum sensing. J Mol Biol 347(2):297–307CrossRefGoogle Scholar
  50. Wang Y, Ma S (2014) Small molecule modulating AHL-based Quorum sensing to attenuate bacterial virulence as promising antimicrobial drugs. Curr Med Chem 21(3):296–311CrossRefGoogle Scholar
  51. Wang J, Wolf RM, Caldwell JW, Kollman PA, Case DA (2004) Development and testing of a general amber force field. J Comput Chem 25(9):1157–1174CrossRefGoogle Scholar
  52. Waters CM, Bassler BL (2006) The Vibrio harveyi quorum-sensing system uses shared regulatory components to discriminate between multiple autoinducers. Genes Dev 20(19):2754–2767CrossRefGoogle Scholar
  53. Yang LW, Eyal E, Bahar I, Kitao A (2009) Principal component analysis of native ensembles of biomolecular structures (PCA_NEST): insights into functional dynamics. Bioinformatics 25:606–614CrossRefGoogle Scholar

Copyright information

© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  1. 1.Department of BiochemistryBose InstituteKolkataIndia
  2. 2.Bioinformatics CentreBose InstituteKolkataIndia
  3. 3.Indian Institute of Science Education and Research Tirupati (IISER-Tirupati)TirupatiIndia
  4. 4.Centre for Computational Biology, The University of KansasLawrenceUSA

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