Department of Pharmaceutical Chemistry, University of California San Francisco
Structural plasticity of staphylococcal nuclease probed by perturbation with pressure and pH
Article first published online: 20 JAN 2011
Copyright © 2010 Wiley-Liss, Inc.
Proteins: Structure, Function, and Bioinformatics
Volume 79, Issue 4, pages 1293–1305, April 2011
How to Cite
Kitahara, R., Hata, K., Maeno, A., Akasaka, K., Chimenti, M. S., Garcia-Moreno E., B., Schroer, M. A., Jeworrek, C., Tolan, M., Winter, R., Roche, J., Roumestand, C., Montet de Guillen, K. and Royer, C. A. (2011), Structural plasticity of staphylococcal nuclease probed by perturbation with pressure and pH. Proteins, 79: 1293–1305. doi: 10.1002/prot.22966
- Issue published online: 8 MAR 2011
- Article first published online: 20 JAN 2011
- Accepted manuscript online: 13 DEC 2010 04:39PM EST
- Manuscript Accepted: 29 NOV 2010
- Manuscript Revised: 1 NOV 2010
- Manuscript Received: 24 AUG 2010
- CAR et CR INSERM, the CNRS, and the ANR PiriBio. Grant Number: 09-455024
- NIH. Grant Numbers: NIH grant GM-061597, NSF grant MCB-0743422
- Deutsche Forschungsgemeinschaft (DFG)
- KA Academic Frontier Program of the Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT);
- 1) Grant-in-Aid for JSPS Fellows and Grant-in-Aid for Scientific Research on Innovative Area of MEXT. Grant Number: MAS would like to acknowledge the Deutsche Forschungsgemeinschaft (DFG TO169/14-1) for financial support. The BL9 and BW4 beamline crews and the machine groups of DELTA and HASYLAB are kindly acknowledged. For their excellent help during the SAXS beamtimes we thank M. Paulus, C. Sternemann, D.C.F. Wieland, Ch.J. Sahle, A. Steffen and S. Tiemeyer.
- protein electrostatics;
- conformational dynamics;
The ionization of internal groups in proteins can trigger conformational change. Despite this being the structural basis of most biological energy transduction, these processes are poorly understood. Small angle X-ray scattering (SAXS) and nuclear magnetic resonance (NMR) spectroscopy experiments at ambient and high hydrostatic pressure were used to examine how the presence and ionization of Lys-66, buried in the hydrophobic core of a stabilized variant of staphylococcal nuclease, affect conformation and dynamics. NMR spectroscopy at atmospheric pressure showed previously that the neutral Lys-66 affects slow conformational fluctuations globally, whereas the effects of the charged form are localized to the region immediately surrounding position 66. Ab initio models from SAXS data suggest that when Lys-66 is charged the protein expands, which is consistent with results from NMR spectroscopy. The application of moderate pressure (<2 kbar) at pH values where Lys-66 is normally neutral at ambient pressure left most of the structure unperturbed but produced significant nonlinear changes in chemical shifts in the helix where Lys-66 is located. Above 2 kbar pressure at these pH values the protein with Lys-66 unfolded cooperatively adopting a relatively compact, albeit random structure according to Kratky analysis of the SAXS data. In contrast, at low pH and high pressure the unfolded state of the variant with Lys-66 is more expanded than that of the reference protein. The combined global and local view of the structural reorganization triggered by ionization of the internal Lys-66 reveals more detectable changes than were previously suggested by NMR spectroscopy at ambient pressure. Proteins 2011. © 2011 Wiley-Liss, Inc.