| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1 Department of Biotechnology, Graduate School of Engineering,
2 Department of Bioinformatic Engineering, Graduate School of Information Science and Technology, and
3 Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan
4 Research Institute for Electronic Science, Hokkaido University, N12W6, Kita-ku, 060-0812, Sapporo, Japan
5 Target Oriented Research for Embroynic Science and Technology (TOREST), Japan Science and Technology Agency (JST), Yamadaoka, Suita, Osaka 565-0871, Japan
6 Department of Pure and Applied Sciences, University of Tokyo, Komaba, Meguro-ku, Tokyo 153-8902, Japan
Reprint requests to: Tetsuya Yomo, Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamada-oka, Suita City, Osaka, 565-0871, Japan; e-mail: yomo{at}bio.eng.osaka-u.ac.jp; fax: 81-6-6879-7428.
Unfolded states of ribonuclease A were used to investigate the effects of macromolecular crowding on macromolecular compactness and protein folding. The extent of protein folding and compactness were measured by circular dichroism spectroscopy, fluorescence correlation spectroscopy, and NMR spectroscopy in the presence of polyethylene glycol (PEG) or Ficoll as the crowding agent. The unfolded state of RNase A in a 2.4 M urea solution at pH 3.0 became native in conformation and compactness by the addition of 35% PEG 20000 or Ficoll 70. In addition, the effects of macromolecular crowding on inert macromolecule compactness were investigated by fluorescence correlation spectroscopy using Fluorescence-labeled PEG as a test macromolecule. The size of Fluorescence-labeled PEG decreased remarkably with an increase in the concentration of PEG 20000 or Ficoll 70. These results show that macromolecules are favored compact conformations in the presence of a high concentration of macromolecules and indicate the importance of a crowded environment for the folding and stabilization of globular proteins. Furthermore, the magnitude of the effects on macromolecular crowding by the different sizes of background molecules was investigated. RNase A and Fluorescence-labeled PEG did not become compact, and had folded conformation by the addition of PEG 200. The effect of the chemical potential on the compaction of a test molecule in relation to the relative sizes of the test and background molecules is also discussed.
Keywords: protein folding; macromolecular crowding; macromolecular compaction; ribonuclease A; polyethylene glycol; circular dichroism; fluorescence correlation spectroscopy; NMR spectroscopy
CiteULike 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  D. Homouz, M. Perham, A. Samiotakis, M. S. Cheung, and P. Wittung-Stafshede Crowded, cell-like environment induces shape changes in aspherical protein PNAS, August 19, 2008; 105(33): 11754 - 11759. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Stagg, S.-Q. Zhang, M. S. Cheung, and P. Wittung-Stafshede Molecular crowding enhances native structure and stability of {alpha}/ protein flavodoxin PNAS, November 27, 2007; 104(48): 18976 - 18981. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Monterroso and A. P. Minton Effect of High Concentration of Inert Cosolutes on the Refolding of an Enzyme: CARBONIC ANHYDRASE B IN SUCROSE AND FICOLL 70 J. Biol. Chem., November 16, 2007; 282(46): 33452 - 33458. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. C. Konopka, I. A. Shkel, S. Cayley, M. T. Record, and J. C. Weisshaar Crowding and confinement effects on protein diffusion in vivo. J. Bacteriol., September 1, 2006; 188(17): 6115 - 6123. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. P. Minton How can biochemical reactions within cells differ from those in test tubes? J. Cell Sci., July 15, 2006; 119(14): 2863 - 2869. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. GUO, J.-E. SHEA, and R S. BERRY The Physics of the Interactions Governing Folding and Association of Proteins Ann. N.Y. Acad. Sci., December 1, 2005; 1066(1): 34 - 53. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Campanini, S. Bologna, F. Cannone, G. Chirico, A. Mozzarelli, and S. Bettati Unfolding of Green Fluorescent Protein mut2 in wet nanoporous silica gels Protein Sci., May 1, 2005; 14(5): 1125 - 1133. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. P. Minton Models for Excluded Volume Interaction between an Unfolded Protein and Rigid Macromolecular Cosolutes: Macromolecular Crowding and Protein Stability Revisited Biophys. J., February 1, 2005; 88(2): 971 - 985. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B.-R. Zhou, Y. Liang, F. Du, Z. Zhou, and J. Chen Mixed Macromolecular Crowding Accelerates the Oxidative Refolding of Reduced, Denatured Lysozyme: IMPLICATIONS FOR PROTEIN FOLDING IN INTRACELLULAR ENVIRONMENTS J. Biol. Chem., December 31, 2004; 279(53): 55109 - 55116. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Zorrilla, G. Rivas, A. U. Acuna, and M. P. Lillo Protein self-association in crowded protein solutions: A time-resolved fluorescence polarization study Protein Sci., November 1, 2004; 13(11): 2960 - 2969. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
