Journal of the Korean Magnetic Resonance Society (한국자기공명학회논문지)

Korean Magnetic Resonance Society (한국자기공명학회)
권호 표지
DOI QR코드

DOI QR Code

Triple isotope-[13C, 15N, 2H] labeling and NMR measurements of the inactive, reduced monomer form of Escherichia coli Hsp33

  • Lee, Yoo-Sup (Department of Biotechnology, Konkuk University) ;
  • Ko, Hyun-Suk (Department of Biotechnology, Konkuk University) ;
  • Ryu, Kyoung-Seok (Division of Magnetic Resonance, Korea Basic Science Institute) ;
  • Jeon, Young-Ho (Division of Magnetic Resonance, Korea Basic Science Institute) ;
  • Won, Hyung-Sik (Department of Biotechnology, Konkuk University)
  • Received : 2010.10.30
  • Accepted : 2010.12.09
  • Published : 2010.12.20
Download PDF
⟨ Previous Next ⟩

Abstract

Hsp33 is a molecular chaperone achieving a holdase activity upon response to a dual stress by heat and oxidation. Despite several crystal structures available, the activation process is not clearly understood, because the structure inactive Hsp33 as its reduced, zinc-bound, monomeric form has not been solved yet. Thus, we initiated structural investigation of the reduced Hsp33 monomer by NMR. In this study, to overcome the high molecular weight (33 kDa), the protein was triply isotope-[$^{13}C$, $^{15}N$, $^2H$]-labeled and its inactive, monomeric state was ensured. 2D-[$^1H$, $^{15}N$]-TROSY and a series of triple resonance spectra could be successfully obtained on a high-field (900 MHz) NMR machine with a cryoprobe. However, under all of the different conditions tested, the number of resonances observed was significantly less than that expected from the amino acid sequence. Thus, a possible contribution of dynamic conformational exchange leading to a line broadening is suggested that might be important for activation process of Hsp33.

Keywords

References

  1. C. Kumsta, U. Jakob, Biochemistry 48, 4666. (2009). https://doi.org/10.1021/bi9003556
  2. U. Jakob, W. Muse, M. Eser, J.C.A. Bardwell, Cell 96, 341. (1999). https://doi.org/10.1016/S0092-8674(00)80547-4
  3. M. Ilbert, J. Horst, S. Ahrens, J. Winter, P.C.F. Graf, H. Lilie, U. Jakob, Nat. Struct. Mol. Biol. 14, 556. (2007). https://doi.org/10.1038/nsmb1244
  4. J. Winter, M. Ilbert, P.C.F. Graf, D. Ozcelik, U. Jakob, Cell 135, 691. (2008). https://doi.org/10.1016/j.cell.2008.09.024
  5. M.W. Akhtar, V. Srinivas, B. Raman, T. Ramakrishna, T. Inobe, K. Maki, M. Arai, K. Kuwajima, C.M. Rao, J. Biol. Chem. 279, 55760. (2004). https://doi.org/10.1074/jbc.M406333200
  6. J.H. Hoffmann, K. Linke, P.C.F. Graf, H. Lilie, U. Jakob, EMBO J. 23, 160. (2004). https://doi.org/10.1038/sj.emboj.7600016
  7. I. Janda, Y. Devedjiev, U. Derewenda, Z. Dauter, J. Bielnicki, D.R. Cooper, P.C.F. Graf, A. Joachimiak, U. Jakob, Z.S. Derewenda, Structure 12, 1901. (2004). https://doi.org/10.1016/j.str.2004.08.003
  8. J. Vijayalakshmi, M.K. Mukhergee, J. Graumann, U. Jakob, M.A. Saper, Structure 9, 367. (2001). https://doi.org/10.1016/S0969-2126(01)00597-4
  9. S.-J. Kim, D.-G. Jeong, S.-W. Chi, J.-S. Lee, S.-E. Ryu, Nat. Struct. Biol. 8, 459. (2001). https://doi.org/10.1038/87639
  10. H.-S. Won, L.Y. Low, R. De Guzman, M. Martinez-Yamout, U. Jakob, H.J. Dyson, J. Mol. Biol. 341, 893. (2004). https://doi.org/10.1016/j.jmb.2004.06.046
  11. L. Jaroszewski, R. Schwarzenbacher, D. McMullan, P. Abdubek, S. Agarwalla, E. Ambing, H. Axelrod, T. Biorac, J.M. Canaves, H. J. Chiu, et al., Proteins 61, 669. (2005). https://doi.org/10.1002/prot.20542
  12. D.-W. Sim, H.-C. Ahn, H.-S. Won, J. Kor. Magn. Reson. Soc. 13, 108. (2009). https://doi.org/10.6564/JKMRS.2009.13.2.108
  13. J.-H. Kim, K.-Y. Lee, S.-J. Park, B.-J. Lee, J. Kor. Magn. Reson. Soc. 14, 45. (2010). https://doi.org/10.6564/JKMRS.2010.14.1.045
  14. P.C.F. Graf, M. Martinez-Yamout, S. VanHaerents, H. Lilie, H.J. Dyson, U. Jakob, J. Bio. Chem. 279, 20529. (2004). https://doi.org/10.1074/jbc.M401764200
  15. B. Odaert, I. Landrieu, K. Dijkstra, G. Schuurman-Wolters, P. Casteels, J.-M. Wieruszeski, D. Inze, R. Scheek, G. Lippens, J. Biol. Chem. 277, 12375. (2002). https://doi.org/10.1074/jbc.M111741200
  16. E. Barbar, Biopolymers 51, 191. (1999). https://doi.org/10.1002/(SICI)1097-0282(1999)51:3<191::AID-BIP3>3.0.CO;2-B
  17. C.M. Cremers, D. Reichmann, J. Hausmann, M. Ilbert, U. Jakob, J. Biol. Chem. 285, 11243. (2010). https://doi.org/10.1074/jbc.M109.084350

Cited by

  1. Verification of the interdomain contact site in the inactive monomer, and the domain-swapped fold in the active dimer of Hsp33 in solution vol.586, pp.4, 2012, https://doi.org/10.1016/j.febslet.2012.01.011
  2. Backbone NMR Assignments of a Prokaryotic Molecular Chaperone, Hsp33 from Escherichia coli vol.16, pp.2, 2012, https://doi.org/10.6564/JKMRS.2012.16.2.172
  3. Backbone NMR Assignments of an Uncharacterized Protein, SF1002 from Shigella flexneri 5a M90T vol.19, pp.1, 2015, https://doi.org/10.6564/JKMRS.2015.19.1.036
  4. Oxidation-Induced Conformational Change of a Prokaryotic Molecular Chaperone, Hsp33, Monitored by Selective Isotope Labeling vol.15, pp.2, 2011, https://doi.org/10.6564/JKMRS.2011.15.2.137
  5. Semi-Empirical Structure Determination of Escherichia coli Hsp33 and Identification of Dynamic Regulatory Elements for the Activation Process vol.427, pp.24, 2015, https://doi.org/10.1016/j.jmb.2015.09.029
  6. Oxidation-induced conformational change of Hsp33, monitored by NMR vol.19, pp.3, 2015, https://doi.org/10.6564/JKMRS.2015.19.3.099