Genetic and Biochemical Dissection of a HisKA Domain Identifies Residues Required Exclusively for Kinase and Phosphatase Activities

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Two-component signal transduction systems, composed of histidine kinases (HK) and response regulators (RR), allow bacteria to respond to diverse environmental stimuli. The HK can control both phosphorylation and subsequent dephosphorylation of its cognate RR. The majority of HKs utilize the HisKA subfamily of dimerization and histidine phosphotransfer (DHp) domains, which contain the phospho-accepting histidine and directly contact the RR. Extensive genetics, biochemistry, and structural biology on several prototypical TCS systems including NtrB-NtrC and EnvZ-OmpR have provided a solid basis for understanding the function of HK–RR signaling. Recently, work on NarX, a HisKA_3 subfamily protein, indicated that two residues in the highly conserved region of the DHp domain are responsible for phosphatase activity. In this study we have carried out both genetic and biochemical analyses on Myxococcus xanthus CrdS, a member of the HisKA subfamily of bacterial HKs. CrdS is required for the regulation of spore formation in response to environmental stress. Following alanine-scanning mutagenesis of the α1 helix of the DHp domain of CrdS, we determined the role for each mutant protein for both kinase and phosphatase activity. Our results indicate that the conserved acidic residue (E372) immediately adjacent to the site of autophosphorylation (H371) is specifically required for kinase activity but not for phosphatase activity. Conversely, we found that the conserved Thr/Asn residue (N375) was required for phosphatase activity but not for kinase activity. We extended our biochemical analyses to two CrdS homologs from M. xanthus, HK1190 and HK4262, as well as Thermotoga maritima HK853. The results were similar for each HisKA family protein where the conserved acidic residue is required for kinase activity while the conserved Thr/Asn residue is required for phosphatase activity. These data are consistent with conserved mechanisms for kinase and phosphatase activities in the broadly occurring HisKA family of sensor kinases in bacteria.

Vyšlo v časopise: Genetic and Biochemical Dissection of a HisKA Domain Identifies Residues Required Exclusively for Kinase and Phosphatase Activities. PLoS Genet 8(11): e32767. doi:10.1371/journal.pgen.1003084
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1003084

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Zdroje

1. ZusmanD, ScottA, YangZ, KirbyJ (2007) Chemosensory pathways, motility and development in Myxococcus xanthus. Nat Rev Microbiol 5: 862–872.

2. UlrichLE, ZhulinIB (2010) The MiST2 database: a comprehensive genomics resource on microbial signal transduction. Nucleic Acids Research 38: D401–D407.

3. KirbyJR, ZusmanDR (2003) Chemosensory regulation of developmental gene expression in Myxococcusxanthus. Proceedings of the National Academy of Sciences of the United States of America 100: 2008–2013.

4. WillettJW, KirbyJR (2011) CrdS and CrdA Comprise a Two-Component System That Is Cooperatively Regulated by the Che3 Chemosensory System in Myxococcus xanthus. mBio 2.

5. ParkH, SahaSK, InouyeM (1998) Two-domain reconstitution of a functional protein histidine kinase. Proceedings of the National Academy of Sciences 95: 6728–6732.

6. CasinoP, RubioV, MarinaA (2010) The mechanism of signal transduction by two-component systems. Current Opinion in Structural Biology 20: 763–771.

7. PazyY, MotalebMA, GuarnieriMT, CharonNW, ZhaoR, et al. (2010) Identical phosphatase mechanisms achieved through distinct modes of binding phosphoprotein substrate. Proceedings of the National Academy of Sciences 107: 1924–1929.

8. ParkS-Y, ChaoX, Gonzalez-BonetG, BeelBD, BilwesAM, et al. (2004) Structure and Function of an Unusual Family of Protein Phosphatases: The Bacterial Chemotaxis Proteins CheC and CheX. Molecular Cell 16: 563–574.

9. ParasharV, MirouzeN, DubnauDA, NeiditchMB (2011) Structural Basis of Response Regulator Dephosphorylation by Rap Phosphatases. PLoS Biol 9: e1000589 doi:10.1371/journal.pbio.1000589

10. DuttaR, YoshidaT, InouyeM (2000) The Critical Role of the Conserved Thr247 Residue in the Functioning of the Osmosensor EnvZ, a Histidine Kinase/Phosphatase, in Escherichia coli. Journal of Biological Chemistry 275: 38645–38653.

11. GutuAD, WayneKJ, ShamL-T, WinklerME (2010) Kinetic Characterization of the WalRKSpn (VicRK) Two-Component System of Streptococcus pneumoniae: Dependence of WalKSpn (VicK) Phosphatase Activity on Its PAS Domain. J Bacteriol 192: 2346–2358.

12. HuynhTN, NoriegaCE, StewartV (2010) Conserved mechanism for sensor phosphatase control of two-component signaling revealed in the nitrate sensor NarX. Proceedings of the National Academy of Sciences 107: 21140–21145.

13. HuynhTN, StewartV (2011) Negative control in two-component signal transduction by transmitter phosphatase activity. Molecular Microbiology 82: 275–286.

14. WolaninPM, WebreDJ, StockJB (2003) Mechanism of Phosphatase Activity in the Chemotaxis Response Regulator CheY†. Biochemistry 42: 14075–14082.

15. FinnRD, MistryJ, TateJ, CoggillP, HegerA, et al. (2010) The Pfam protein families database. Nucleic Acids Research 38: D211–D222.

16. ZhuY, QinL, YoshidaT, InouyeM (2000) Phosphatase activity of histidine kinase EnvZ without kinase catalytic domain. Proceedings of the National Academy of Sciences 97: 7808–7813.

17. KramerG, WeissV (1999) Functional dissection of the transmitter module of the histidine kinase NtrB in Escherichia coli. Proceedings of the National Academy of Sciences 96: 604–609.

18. SkarpholK, WaukauJ, ForstSA (1997) Role of His243 in the phosphatase activity of EnvZ in Escherichia coli. Journal of Bacteriology 179: 1413–1416.

19. HsingW, RussoFD, BerndKK, SilhavyTJ (1998) Mutations That Alter the Kinase and Phosphatase Activities of the Two-Component Sensor EnvZ. Journal of Bacteriology 180: 4538–4546.

20. PioszakAA, NinfaAJ (2003) Genetic and Biochemical Analysis of Phosphatase Activity of Escherichia coli NRII (NtrB) and Its Regulation by the PII Signal Transduction Protein. Journal of Bacteriology 185: 1299–1315.

21. CapraEJ, PerchukBS, LubinEA, AshenbergO, SkerkerJM, et al. (2010) Systematic Dissection and Trajectory-Scanning Mutagenesis of the Molecular Interface That Ensures Specificity of Two-Component Signaling Pathways. PLoS Genet 6: e1001220 doi:10.1371/journal.pgen.1001220

22. QinL, CaiS, ZhuY, InouyeM (2003) Cysteine-Scanning Analysis of the Dimerization Domain of EnvZ, an Osmosensing Histidine Kinase. Journal of Bacteriology 185: 3429–3435.

23. CasinoP, RubioV, MarinaA (2009) Structural Insight into Partner Specificity and Phosphoryl Transfer in Two-Component Signal Transduction. Cell 139: 325–336.

24. SkerkerJM, PerchukBS, SiryapornA, LubinEA, AshenbergO, et al. (2008) Rewiring the Specificity of Two-Component Signal Transduction Systems. Cell 133: 1043–1054.

25. YamadaM, MakinoK, AmemuraM, ShinagawaH, NakataA (1989) Regulation of the phosphate regulon of Escherichia coli: analysis of mutant phoB and phoR genes causing different phenotypes. Journal of Bacteriology 171: 5601–5606.

26. MarinaA, WaldburgerCD, HendricksonWA (2005) Structure of the entire cytoplasmic portion of a sensor histidine-kinase protein. Embo J 24: 4247–4259.

27. TomomoriC, TanakaT, DuttaR, ParkH, SahaSK, et al. (1999) Solution structure of the homodimeric core domain of Escherichia coli histidine kinase EnvZ. Nat Struct Mol Biol 6: 729–734.

28. Grebe TW, Stock JB (1999) The Histidine Protein Kinase Superfamily. In: Poole RK, editor. Advances in Microbial Physiology: Academic Press. pp. 139–227.

29. CamachoC, CoulourisG, AvagyanV, MaN, PapadopoulosJ, et al. (2009) BLAST+: architecture and applications. BMC Bioinformatics 10: 421.

30. CrooksGE, HonG, ChandoniaJ-M, BrennerSE (2004) WebLogo: A Sequence Logo Generator. Genome Research 14: 1188–1190.

31. ChenYE, TsokosCG, BiondiEG, PerchukBS, LaubMT (2009) Dynamics of Two Phosphorelays Controlling Cell Cycle Progression in Caulobacter crescentus. Journal of Bacteriology 191: 7417–7429.

32. HsingW, SilhavyTJ (1997) Function of conserved histidine-243 in phosphatase activity of EnvZ, the sensor for porin osmoregulation in Escherichia coli. Journal of Bacteriology 179: 3729–3735.

33. XuQ, BlackWP, WardSM, YangZ (2005) Nitrate-Dependent Activation of the Dif Signaling Pathway of Myxococcus xanthus Mediated by a NarX-DifA Interspecies Chimera. J Bacteriol 187: 6410–6418.

34. WuS, KaiserD (1997) Regulation of expression of the pilA gene in Myxococcus xanthus. J Bacteriol 179: 7748–7758.

35. MagriniV, CreightonC, YouderianP (1999) Site-Specific Recombination of Temperate Myxococcus xanthus Phage Mx8: Genetic Elements Required for Integration. J Bacteriol 181: 4050–4061.

36. MatrouleJ-Y, LamH, BurnetteDT, Jacobs-WagnerC (2004) Cytokinesis Monitoring during Development: Rapid Pole-to-Pole Shuttling of a Signaling Protein by Localized Kinase and Phosphatase in Caulobacter. Cell 118: 579–590.

37. AtkinsonMR, NinfaAJ (1993) Mutational analysis of the bacterial signal-transducing protein kinase/phosphatase nitrogen regulator II (NRII or NtrB). Journal of Bacteriology 175: 7016–7023.

38. RuthES (2010) Auxiliary phosphatases in two-component signal transduction. Current Opinion in Microbiology 13: 177–183.

39. NakanoMM, ZhuY (2001) Involvement of ResE Phosphatase Activity in Down-Regulation of ResD-Controlled Genes in Bacillus subtilis during Aerobic Growth. Journal of Bacteriology 183: 1938–1944.

40. RaivioTL, SilhavyTJ (1997) Transduction of envelope stress in Escherichia coli by the Cpx two-component system. Journal of Bacteriology 179: 7724–7733.

41. KroosL (2007) The Bacillus and Myxococcus Developmental Networks and Their Transcriptional Regulators. Annual Review of Genetics 41: 13–39.

42. KelleyLA, SternbergMJE (2009) Protein structure prediction on the Web: a case study using the Phyre server. Nat Protocols 4: 363–371.

43. BlackhartBD, ZusmanDR (1985) “Frizzy” genes of Myxococcus xanthus are involved in control of frequency of reversal of gliding motility. Proceedings of the National Academy of Sciences 82: 8767–8770.

44. LeeB, SchrammA, JagadeesanS, HiggsPI (2010) Two-component systems and regulation of developmental progression in Myxococcus xanthus. Methods Enzymol 471: 253–278.