Bulletin of the Korean Chemical Society

  • 제33권9호
  • /
  • Pages.2981-2984
  • /
  • 2012
  • /
  • 0253-2964(pISSN)
  • /
  • 1229-5949(eISSN)
대한화학회 (Korean Chemical Society)
권호 표지
DOI QR코드

DOI QR Code

Rapid Mapping of Active Site of KSI by Paramagnetic NMR

  • Joe, Yong-Nam (Department of Chemistry, Pohang University of Science and Technology) ;
  • Cha, Hyung-Jin (Department of Life Science, Pohang University of Science and Technology) ;
  • Lee, Hyeong-Ju (Department of Chemistry, Pohang University of Science and Technology) ;
  • Choi, Kwan-Yong (Department of Life Science, Pohang University of Science and Technology) ;
  • Lee, Hee-Cheon (Department of Chemistry, Pohang University of Science and Technology)
  • 투고 : 2012.06.02
  • 심사 : 2012.06.12
  • 발행 : 2012.09.20
PDF 다운로드
⟨ 이전 논문 다음 논문 ⟩

초록

Active site mapping has been done for ${\Delta}^5$-3-ketosteroid isomerase (KSI) by analyses of paramagnetic effect on $^1H-^{15}N$ HSQC spectra using 4-hydroxyl-2,2,6,6-tetramethylpiperidinyl-1-oxy (HyTEMPO) and an intermediate analog (equilenin). Our result revealed that residues in hydrophobic cavity of KSI, particularly active site region, mainly experienced a high line-broadening effect of NMR signal with HyTEMPO, while they experienced full recovery of a lineshape upon the addition of equilenin. The mapped region was very similar to the active site of KSI as described by the crystal structure. These observations indicate that a combined use of paramagnetic reagent and substrate (or analog) could rapidly identify the residues in potential active site of KSI, and can be applied to the analysis of both active site and function in unknown protein.

키워드

참고문헌

  1. Koshland, D. E. Proc. Natl. Acad Sci. USA 1958, 44, 98. https://doi.org/10.1073/pnas.44.2.98
  2. Berzofsky, J. A. Science 1985, 229, 932. https://doi.org/10.1126/science.2410982
  3. Chothia, C.; Janin, J. Nature 1975, 256, 705. https://doi.org/10.1038/256705a0
  4. Lo Conte, L.; Chothia, C.; Janin, J. J. Mol. Biol. 1999, 285, 2177. https://doi.org/10.1006/jmbi.1998.2439
  5. Li, N.; Sun, Z.; Jiang, F. BMC Bioinformatics 2008, 9, 553. https://doi.org/10.1186/1471-2105-9-553
  6. Blow, D. B.; Brick, P.; Collyer, C. A.; Goldberg, J. D.; Smart, O. Phil. Trans. R. Soc. Lond. A 1992, 340, 311. https://doi.org/10.1098/rsta.1992.0069
  7. Smiley, R. D.; Hammes, G. G. Chem. Rev. 2006, 106, 3080. https://doi.org/10.1021/cr0502955
  8. Barth, A.; Zscherp, C. FEBS Lett. 2000, 477, 151. https://doi.org/10.1016/S0014-5793(00)01782-8
  9. Berman, H. M.; Westbrook, J.; Feng, Z.; Gilliland, G.; Bhat, T. N.; Weissig, H.; Shindyalov, I. N.; Bourne, P. E. Nucleic. Acids Res. 2000, 28, 235. https://doi.org/10.1093/nar/28.1.235
  10. Petros, A. M.; Mueller, L.; Kopple, K. D. Biochemistry 1990, 29, 10041. https://doi.org/10.1021/bi00495a005
  11. Moore, C. D.; Lecomte, J. T. Biochemistry 1993, 32, 199. https://doi.org/10.1021/bi00052a026
  12. Batzold, F. H.; Benson, A. M.; Robinson, C. H.; Talalay, P. Adv. Enzyme Regul. 1976, 14, 243. https://doi.org/10.1016/0065-2571(76)90016-9
  13. Hawkinson, D. C.; Eames, T. C.; Pollack, R. M. Biochemistry 1991, 30, 10849. https://doi.org/10.1021/bi00109a007
  14. Hawkinson, D. C.; Pollack, R. M.; Ambulos, N. P., Jr. Biochemistry 1994, 33, 12172. https://doi.org/10.1021/bi00206a021
  15. Wu, Z. R.; Ebrahimian, S.; Zawrotny, M. E.; Thornburg, L. D.; Perez-Alvarado, G. C.; Brothers, P.; Pollack, R. M.; Summers, M. F. Science 1997, 276, 415. https://doi.org/10.1126/science.276.5311.415
  16. 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.; Oh, B. H. Biochemistry 1997, 36, 14030. https://doi.org/10.1021/bi971546+
  17. Kraut, D. A.; Sigala, P. A.; Pybus, B.; Liu, C. W.; Ringe, D.; Petsko, G. A.; Herschlag, D. PLoS Biol. 2006, 4, e99. https://doi.org/10.1371/journal.pbio.0040099
  18. Nam, G. H.; Kim, D.; Ha, N.; Jang, D. S.; Yun, Y. S.; Hong, B. H.; Oh, B.; Choi, K. Y. J. Biochem. 2003, 134, 101. https://doi.org/10.1093/jb/mvg117
  19. Choi, G.; Ha, N. C.; Kim, S. W.; Kim, D. H.; Park, S.; Oh, B. H.; Choi, K. Y. Biochemistry 2000, 39, 903. https://doi.org/10.1021/bi991579k
  20. Wishart, D. S.; Biqam, C. G.; Yao, J.; Abildqaard, F.; Dyson, H. J.; Oldfield, E.; Markley, J. L.; Sykes, B. D. J. Biomol. NMR 1995, 6, 135.
  21. Delaqlio, F.; Grzesiek, S.; Vuister, G. W.; Zhu, G.; Pfeifer, J.; Bax, A. J. Biomol. NMR 1995, 6, 277.
  22. Goddard, T. D.; Kneller, D. G. Sparky3, University of california, San Fransisco.
  23. Cornilescu, G.; Delaglio, F.; Bax, A. J. Biomol. NMR 1999, 13, 289. https://doi.org/10.1023/A:1008392405740
  24. Lee, H. J.; Jang, D. S.; Cha, H. J.; Moon, H. S.; Hong, B. H.; Choi, K. Y.; Lee, H. C. J. Biochem. 2008, 144, 215. https://doi.org/10.1093/jb/mvn058
  25. Esposito, G.; Lesk, A. M.; Molinari, H.; Motta, A.; Niccolai, N.; Pastore, A. J. Mol. Biol. 1992, 224, 659. https://doi.org/10.1016/0022-2836(92)90551-T
  26. Cocco, M. J.; Lecomte, J. T. Protein Sci. 1994, 3, 267.
  27. Molinari, H.; Esposito, G.; Raqona, L.; Peqna, M.; Niccolai, N.; Brunne, R. M.; Lesk, R. M.; Zetta, L. Biophys. J. 1997, 73, 382. https://doi.org/10.1016/S0006-3495(97)78078-0
  28. Yang, X.; Chasteen, N. D. Biophys. J. 1996, 71, 1587. https://doi.org/10.1016/S0006-3495(96)79361-X
  29. Storch, E. M.; Daggett, V. Biochemistry 1995, 34, 9682. https://doi.org/10.1021/bi00030a005
  30. Cho, H. S.; Ha, N. C.; Choi, G.; Kim, H. J.; Lee, D.; Oh, K. S.; Kim, K. S.; Lee, W. T.; Choi, K. Y.; Oh, B. H. J. Biol. Chem. 1999, 274, 32863. https://doi.org/10.1074/jbc.274.46.32863
  31. Pollack, R. M. Bioorg. Chem. 2004, 32, 341. https://doi.org/10.1016/j.bioorg.2004.06.005