Molecules and Cells

Korean Society for Molecular and Cellular Biology (한국분자세포생물학회)
권호 표지
DOI QR코드

DOI QR Code

Contribution of a Low-Barrier Hydrogen Bond to Catalysis Is Not Significant in Ketosteroid Isomerase

  • Jang, Do Soo (Department of Life Sciences, Pohang University of Science and Technology) ;
  • Choi, Gildon (Korea Research Institute of Chemical Technology) ;
  • Cha, Hyung Jin (Department of Life Sciences, Pohang University of Science and Technology) ;
  • Shin, Sejeong (Department of Cell Biology, Harvard Medical School) ;
  • Hong, Bee Hak (Department of Life Sciences, Pohang University of Science and Technology) ;
  • Lee, Hyeong Ju (Department of Chemistry, Pohang University of Science and Technology) ;
  • Lee, Hee Cheon (Department of Chemistry, Pohang University of Science and Technology) ;
  • Choi, Kwan Yong (Department of Life Sciences, Pohang University of Science and Technology)
  • Received : 2014.10.02
  • Accepted : 2015.02.16
  • Published : 2015.05.31
Download PDF
⟨ Previous Next ⟩

Abstract

Low-barrier hydrogen bonds (LBHBs) have been proposed to have important influences on the enormous reaction rate increases achieved by many enzymes. ${\Delta}^5$-3-ketosteroi isomerase (KSI) catalyzes the allylic isomerization of ${\Delta}^5$-3-ketosteroid to its conjugated ${\Delta}^4$-isomers at a rate that approache the diffusion limit. Tyr14, a catalytic residue of KSI, has been hypothesized to form an LBHB with the oxyanion of a dienolate steroid intermediate generated during the catalysis. The unusual chemical shift of a proton at 16.8 ppm in the nuclear magnetic resonance spectrum has been attributed to an LBHB between Tyr14 $O{\eta}$ and C3-O of equilenin an intermediate analogue, in the active site of D38N KSI. This shift in the spectrum was not observed in Y30F/Y55F/D38N and Y30F/Y55F/Y115F/D38N mutant KSIs when each mutant was complexed with equilenin, suggesting that Tyr14 could not form LBHB with the intermediate analogue in these mutant KSIs. The crystal structure of Y30F/Y55F/Y115F/D38N-equilenin complex revealed that the distance between Tyr14 $O{\eta}$ and C3-O of the bound steroi was within a direct hydrogen bond. The conversion of LBHB to an ordinary hydrogen bond in the mutant KSI reduced the binding affinity for the steroid inhibitors by a factor of 8.1-11. In addition, the absence of LBHB reduced the catalytic activity by only a factor of 1.7-2. These results suggest that the amount of stabilization energy of the reaction intermediate provided by LBHB is small compared with that provided by an ordinary hydrogen bond in KSI.

Keywords

References

  1. Afonine, P.V., Grosse-Kunstleve, R.W., Echols, N., Headd, J.J., Moriarty, N.W., Mustyakimov, M., Terwilliger, T.C., Urzhumtsev, A., Zwart, P.H., and Adams, P.D. (2012). Towards automated crystallographic structure refinement with phenix.refine. Acta Crystallogr. D Biol. Crystallogr. 68, 352-367.
  2. Ash, E.L., Sudmeier, J.L., De Fabo, E.C., and Bachovchin, W.W. (1997). A low-barrier hydrogen bond in the catalytic triad of serine proteases? Theory versus experiment. Science 278, 1128-1132. https://doi.org/10.1126/science.278.5340.1128
  3. Cha, H.J., Jang, D.S., Kim, Y.G., Hong, B.H., Woo, J.S., Kim, K.T., and Choi, K.Y. (2013). Rescue of deleterious mutations by the compensatory Y30F mutation in ketosteroid isomerase. Mol. Cells 36, 39-46. https://doi.org/10.1007/s10059-013-0013-1
  4. Cha, H.J., Jeong, J.H., Rojviriya, C., and Kim, Y.G. (2014). Structure of Putrescine Aminotransferase from Escherichia coli Provides Insights into the Substrate Specificity among Class III Aminotransferases. PLoS One 9, e113212. https://doi.org/10.1371/journal.pone.0113212
  5. Cho, H.S., Choi, G., Choi, K.Y., and Oh, B.H. (1998). Crystal structure and enzyme mechanism of Delta 5-3-ketosteroid isomerase from Pseudomonas testosteroni. Biochemistry 37, 8325-8330. https://doi.org/10.1021/bi9801614
  6. Cho, H.S., Ha, N.C., Choi, G., Kim, H.J., Lee, D., Oh, K.S., Kim, K.S., Lee, W., Choi, K.Y., and Oh, B.H. (1999). Crystal structure of delta(5)-3-ketosteroid isomerase from Pseudomonas testosteroni in complex with equilenin settles the correct hydrogen bonding scheme for transition state stabilization. J. Biol. Chem. 274, 32863-32868. https://doi.org/10.1074/jbc.274.46.32863
  7. Cleland, W.W., and Kreevoy, M.M. (1994). Low-barrier hydrogen bonds and enzymic catalysis. Science 264, 1887-1890. https://doi.org/10.1126/science.8009219
  8. Cleland, W.W., Frey, P.A., and Gerlt, J.A. (1998). The low barrier hydrogen bond in enzymatic catalysis. J. Biol. Chem. 273, 25529-25532. https://doi.org/10.1074/jbc.273.40.25529
  9. Emsley, P., Lohkamp, B., Scott, W.G., and Cowtan, K. (2010). Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486-501. https://doi.org/10.1107/S0907444910007493
  10. Frey, P.A., Whitt, S.A., and Tobin, J.B. (1994). A low-barrier hydrogen bond in the catalytic triad of serine proteases. Science 264, 1927-1930. https://doi.org/10.1126/science.7661899
  11. Fuhrmann, C.N., Daugherty, M.D., and Agard, D.A. (2006). Subangstrom crystallography reveals that short ionic hydrogen bonds, and not a His-Asp low-barrier hydrogen bond, stabilize the transition state in serine protease catalysis. J. Am. Chem. Soc. 128, 9086-9102. https://doi.org/10.1021/ja057721o
  12. Gerlt, J.A., Kozarich, J.W., Kenyon, G.L., and Gassman, P.G. (1991). Electrophilic catalysis can explain the unexpected acidity of carbon acids in enzyme-catalyzed reactions. J. Am. Chem. Soc. 113, 9667-9669. https://doi.org/10.1021/ja00025a039
  13. Gerlt, J.A., and Gassman, P.G. (1993). Understanding the rates of certain enzyme-catalyzed reactions: proton abstraction from carbon acids, acyl-transfer reactions, and displacement reactions of phosphodiesters. Biochemistry 32, 11943-11952. https://doi.org/10.1021/bi00096a001
  14. Ha, N.C., Kim, M.S., Lee, W., Choi, K.Y., and Oh, B.H. (2000). Detection of large pKa perturbations of an inhibitor and a catalytic group at an enzyme active site, a mechanistic basis for catalytic power of many enzymes. J. Biol. Chem. 275, 41100-41106. https://doi.org/10.1074/jbc.M007561200
  15. Ha, N.C., Choi, G., Choi, K.Y., and Oh, B.H. (2001). Structure and enzymology of Delta5-3-ketosteroid isomerase. Curr. Opin. Struct. Biol. 11, 674-678. https://doi.org/10.1016/S0959-440X(01)00268-8
  16. Jang, D.S., Cha, H.J., Cha, S.S., Hong, B.H., Ha, N.C., Lee, J.Y., Oh, B.H., Lee, H.S., and Choi, K.Y. (2004). Structural doublemutant cycle analysis of a hydrogen bond network in ketosteroid isomerase from Pseudomonas putida biotype B. Biochem. J. 382, 967-973. https://doi.org/10.1042/BJ20031871
  17. Jang, D.S., Lee, H.J., Lee, B., Hong, B.H., Cha, H.J., Yoon, J., Lim, K., Yoon, Y.J., Kim, J., Ree, M., et al. (2006). Detection of an intermediate during the unfolding process of the dimeric ketosteroid isomerase. FEBS Lett. 580, 4166-4171. https://doi.org/10.1016/j.febslet.2006.06.069
  18. Kim, S.W., and Choi, K.Y. (1995). Identification of active site residues by site-directed mutagenesis of delta 5-3-ketosteroid isomerase from Pseudomonas putida biotype B. J. Bacteriol. 177, 2602-2605. https://doi.org/10.1128/jb.177.9.2602-2605.1995
  19. Kim, S.W., Cha, S.S., Cho, H.S., Kim, J.S., Ha, N.C., Cho, M.J., Joo, S., Kim, K.K., Choi, K.Y., and Oh, B.H. (1997a). High-resolution crystal structures of delta5-3-ketosteroid isomerase with and without a reaction intermediate analogue. Biochemistry 36, 14030-14036. https://doi.org/10.1021/bi971546+
  20. Kim, S.W., Joo, S., Choi, G., Cho, H.S., Oh, B.H., and Choi, K.Y. (1997b). Mutational analysis of the three cysteines and active-site aspartic acid 103 of ketosteroid isomerase from Pseudomonas putida biotype B. J. Bacteriol. 179, 7742-7747. https://doi.org/10.1128/jb.179.24.7742-7747.1997
  21. Kim, D.H., Nam, G.H., Jang, D.S., Choi, G., Joo, S., Kim, J.S., Oh, B.H., and Choi, K.Y. (1999). Roles of active site aromatic residues in catalysis by ketosteroid isomerase from Pseudomonas putida biotype B. Biochemistry 38, 13810-13819. https://doi.org/10.1021/bi991040m
  22. Kim, D.H., Jang, D.S., Nam, G.H., Choi, G., Kim, J.S., Ha, N.C., Kim, M.S., Oh, B.H., and Choi, K.Y. (2000). Contribution of the hydrogen-bond network involving a tyrosine triad in the active site to the structure and function of a highly proficient ketosteroid isomerase from Pseudomonas putida biotype B. Biochemistry 39, 4581-4589. https://doi.org/10.1021/bi992119u
  23. Kuliopulos, A., Mildvan, A.S., Shortle, D., and Talalay, P. (1989). Kinetic and ultraviolet spectroscopic studies of active-site mutants of delta 5-3-ketosteroid isomerase. Biochemistry 28, 149-159. https://doi.org/10.1021/bi00427a022
  24. Otwinowski, Z., and Minor, W. (1997). Processing of X-ray diffraction data collected in oscillation mode. Meth. Enzymol. 276, 307-326. https://doi.org/10.1016/S0076-6879(97)76066-X
  25. Perrin, C.L. (2010). Are short, low-barrier hydrogen bonds unusually strong? Acc. Chem. Res. 43, 1550-1557. https://doi.org/10.1021/ar100097j
  26. Petrounia, I.P., and Pollack, R.M. (1998). Substituent effects on the binding of phenols to the D38N mutant of 3-oxo-delta5-steroid isomerase. A probe for the nature of hydrogen bonding to the intermediate. Biochemistry 37, 700-705. https://doi.org/10.1021/bi972262s
  27. Plateau, P., and Gueron, M. (1982). Exchangeable proton NMR without base-line distorsion, using new strong-pulse sequences. J. Am. Chem. Soc. 104, 7310-7311. https://doi.org/10.1021/ja00389a067
  28. Pollack, R.M. (2004). Enzymatic mechanisms for catalysis of enolization: ketosteroid isomerase. Bioorg. Chem. 32, 341-353. https://doi.org/10.1016/j.bioorg.2004.06.005
  29. Pollack, R.M., Thornburg, L.D., Wu, Z.R., and Summers, M.F. (1999). Mechanistic insights from the three-dimensional structure of 3-oxo-Delta(5)-steroid isomerase. Arch. Biochem. Biophys. 370, 9-15. https://doi.org/10.1006/abbi.1999.1384
  30. Schutz, C.N., and Warshel, A. (2004). The low barrier hydrogen bond (LBHB) proposal revisited: the case of the Asp... His pair in serine proteases. Proteins 55, 711-723. https://doi.org/10.1002/prot.20096
  31. Shan, S.O., Loh, S., and Herschlag, D. (1996). The energetics of hydrogen bonds in model systems: implications for enzymatic catalysis. Science 272, 97-101. https://doi.org/10.1126/science.272.5258.97
  32. Stratton, J.R., Pelton, J.G., and Kirsch, J.F. (2001). A novel engineered subtilisin BPN' lacking a low-barrier hydrogen bond in the catalytic triad. Biochemistry 40, 10411-10416. https://doi.org/10.1021/bi015542n
  33. Usher, K.C., Remington, S.J., Martin, D.P., and Drueckhammer, D.G. (1994). A very short hydrogen bond provides only moderate stabilization of an enzyme-inhibitor complex of citrate synthase. Biochemistry 33, 7753-7759. https://doi.org/10.1021/bi00191a002
  34. Wang, Z., Luecke, H., Yao, N., and Quiocho, F.A. (1997). A low energy short hydrogen bond in very high resolution structures of protein receptor--phosphate complexes. Nat. Struct. Biol. 4, 519-522. https://doi.org/10.1038/nsb0797-519
  35. Warshel, A., and Papazyan, A. (1996). Energy considerations show that low-barrier hydrogen bonds do not offer a catalytic advantage over ordinary hydrogen bonds. Proc. Natl. Acad. Sci. USA 93, 13665-13670. https://doi.org/10.1073/pnas.93.24.13665
  36. Wu, Z.R., Ebrahimian, S., Zawrotny, M.E., Thornburg, L.D., Perez- Alvarado, G.C., Brothers, P., Pollack, R.M., and Summers, M.F. (1997). Solution structure of 3-oxo-delta5-steroid isomerase. Science 276, 415-418. https://doi.org/10.1126/science.276.5311.415
  37. Xue, L.A., Talalay, P., and Mildvan, A.S. (1991). Studies of the catalytic mechanism of an active-site mutant (Y14F) of delta 5-3- ketosteroid isomerase by kinetic deuterium isotope effects. Biochemistry 30, 10858-10865. https://doi.org/10.1021/bi00109a008
  38. Zhang, C. (2007). Low-barrier hydrogen bond plays key role in active photosystem II--a new model for photosynthetic water oxidation. Biochim. Biophys. Acta. 1767, 493-499. https://doi.org/10.1016/j.bbabio.2006.12.008
  39. Zhao, Q., Li, Y.K., Mildvan, A.S., and Talalay, P. (1995). Ultraviolet spectroscopic evidence for decreased motion of the active site tyrosine residue of delta 5-3-ketosteroid isomerase by steroid binding. Biochemistry 34, 6562-6572. https://doi.org/10.1021/bi00019a038
  40. Zhao, Q., Abeygunawardana, C., Talalay, P., and Mildvan, A.S. (1996). NMR evidence for the participation of a low-barrier hydrogen bond in the mechanism of delta 5-3-ketosteroid isomerase. Proc. Natl. Acad. Sci. USA 93, 8220-8224. https://doi.org/10.1073/pnas.93.16.8220
  41. Zhao, Q., Abeygunawardana, C., Gittis, A.G., and Mildvan, A.S. (1997). Hydrogen bonding at the active site of delta 5-3- ketosteroid isomerase. Biochemistry 36, 14616-14626. https://doi.org/10.1021/bi971549m
  42. Zhao, L., Liao, H., and Tsai, M.D. (2004). The catalytic role of aspartate in a short strong hydrogen bond of the Asp274-His32 catalytic dyad in phosphatidylinositol-specific phospholipase C can be substituted by a chloride ion. J. Biol. Chem. 279, 31995-32000. https://doi.org/10.1074/jbc.M404184200

Cited by

  1. Role of conserved Met112 residue in the catalytic activity and stability of ketosteroid isomerase vol.1864, pp.10, 2016, https://doi.org/10.1016/j.bbapap.2016.06.016
  2. Perturbation of Short Hydrogen Bonds in Photoactive Yellow Protein via Noncanonical Amino Acid Incorporation vol.123, pp.23, 2015, https://doi.org/10.1021/acs.jpcb.9b01571
  3. Genome-Wide Transcriptome Profiling Provides Insight on Cholesterol and Lithocholate Degradation Mechanisms in Nocardioides simplex VKM Ac-2033D vol.11, pp.10, 2015, https://doi.org/10.3390/genes11101229
  4. On the Case of the Misplaced Hydrogens vol.22, pp.2, 2015, https://doi.org/10.1002/cbic.202000376