Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 2001 Feb 15;354(Pt 1):79–87. doi: 10.1042/0264-6021:3540079

The molecular chaperone alpha-crystallin is in kinetic competition with aggregation to stabilize a monomeric molten-globule form of alpha-lactalbumin.

R A Lindner 1, T M Treweek 1, J A Carver 1
PMCID: PMC1221631  PMID: 11171082

Abstract

In vivo, alpha-crystallin and other small heat-shock proteins (sHsps) act as molecular chaperones to prevent the precipitation of 'substrate' proteins under stress conditions through the formation of a soluble sHsp-substrate complex. Using a range of different salt conditions, the rate and extent of precipitation of reduced alpha-lactalbumin have been altered. The interaction of alpha-crystallin with reduced alpha-lactalbumin under these various salt conditions was then studied using a range of spectroscopic techniques. Under conditions of low salt, alpha-lactalbumin aggregates but does not precipitate. alpha-Crystallin is able to prevent this aggregation, initially by stabilization of a monomeric molten-globule species of alpha-lactalbumin. It is proposed that this stabilization occurs through weak transient interactions between alpha-crystallin and alpha-lactalbumin. Eventually a stable, soluble high-molecular-mass complex is formed between the two proteins. Thus it appears that a tendency for alpha-lactalbumin to aggregate (but not necessarily precipitate) is the essential requirement for alpha-crystallin-alpha-lactalbumin interaction. In other words, alpha-crystallin interacts with a non-aggregated form of the substrate to prevent aggregation. The rate of precipitation of alpha-lactalbumin is increased significantly in the presence of Na2SO4 compared with NaCl. However, in the former case, alpha-crystallin is unable to prevent this aggregation and precipitation except in the presence of a large excess of alpha-crystallin, i.e. at mass ratios more than 10 times greater than in the presence of NaCl. It is concluded that a kinetic competition exists between aggregation and interaction of unfolding proteins with alpha-crystallin.

Full Text

The Full Text of this article is available as a PDF (193.9 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Balbach J., Forge V., van Nuland N. A., Winder S. L., Hore P. J., Dobson C. M. Following protein folding in real time using NMR spectroscopy. Nat Struct Biol. 1995 Oct;2(10):865–870. doi: 10.1038/nsb1095-865. [DOI] [PubMed] [Google Scholar]
  2. Carver J. A., Aquilina J. A., Truscott R. J., Ralston G. B. Identification by 1H NMR spectroscopy of flexible C-terminal extensions in bovine lens alpha-crystallin. FEBS Lett. 1992 Oct 19;311(2):143–149. doi: 10.1016/0014-5793(92)81386-z. [DOI] [PubMed] [Google Scholar]
  3. Carver J. A., Guerreiro N., Nicholls K. A., Truscott R. J. On the interaction of alpha-crystallin with unfolded proteins. Biochim Biophys Acta. 1995 Oct 25;1252(2):251–260. doi: 10.1016/0167-4838(95)00146-l. [DOI] [PubMed] [Google Scholar]
  4. Collins K. D., Washabaugh M. W. The Hofmeister effect and the behaviour of water at interfaces. Q Rev Biophys. 1985 Nov;18(4):323–422. doi: 10.1017/s0033583500005369. [DOI] [PubMed] [Google Scholar]
  5. Das K. P., Surewicz W. K. On the substrate specificity of alpha-crystallin as a molecular chaperone. Biochem J. 1995 Oct 15;311(Pt 2):367–370. doi: 10.1042/bj3110367. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Das K. P., Surewicz W. K. Temperature-induced exposure of hydrophobic surfaces and its effect on the chaperone activity of alpha-crystallin. FEBS Lett. 1995 Aug 7;369(2-3):321–325. doi: 10.1016/0014-5793(95)00775-5. [DOI] [PubMed] [Google Scholar]
  7. Ehrnsperger M., Gräber S., Gaestel M., Buchner J. Binding of non-native protein to Hsp25 during heat shock creates a reservoir of folding intermediates for reactivation. EMBO J. 1997 Jan 15;16(2):221–229. doi: 10.1093/emboj/16.2.221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ewbank J. J., Creighton T. E. The molten globule protein conformation probed by disulphide bonds. Nature. 1991 Apr 11;350(6318):518–520. doi: 10.1038/350518a0. [DOI] [PubMed] [Google Scholar]
  9. Hayer-Hartl M. K., Ewbank J. J., Creighton T. E., Hartl F. U. Conformational specificity of the chaperonin GroEL for the compact folding intermediates of alpha-lactalbumin. EMBO J. 1994 Jul 1;13(13):3192–3202. doi: 10.1002/j.1460-2075.1994.tb06618.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Horwitz J., Huang Q. L., Ding L., Bova M. P. Lens alpha-crystallin: chaperone-like properties. Methods Enzymol. 1998;290:365–383. doi: 10.1016/s0076-6879(98)90032-5. [DOI] [PubMed] [Google Scholar]
  11. Jaenicke R. Folding and association versus misfolding and aggregation of proteins. Philos Trans R Soc Lond B Biol Sci. 1995 Apr 29;348(1323):97–105. doi: 10.1098/rstb.1995.0050. [DOI] [PubMed] [Google Scholar]
  12. Jakob U., Gaestel M., Engel K., Buchner J. Small heat shock proteins are molecular chaperones. J Biol Chem. 1993 Jan 25;268(3):1517–1520. [PubMed] [Google Scholar]
  13. KRONMAN M. J., ANDREOTTI R., VITOLS R. INTER- AND INTRAMOLECULAR INTERACTIONS OF ALPHA-LACTALBUMIN. II. AGGREGATION REACTIONS AT ACID PH. Biochemistry. 1964 Aug;3:1152–1160. doi: 10.1021/bi00896a025. [DOI] [PubMed] [Google Scholar]
  14. Kiefhaber T., Rudolph R., Kohler H. H., Buchner J. Protein aggregation in vitro and in vivo: a quantitative model of the kinetic competition between folding and aggregation. Biotechnology (N Y) 1991 Sep;9(9):825–829. doi: 10.1038/nbt0991-825. [DOI] [PubMed] [Google Scholar]
  15. Kuwajima K. The molten globule state of alpha-lactalbumin. FASEB J. 1996 Jan;10(1):102–109. doi: 10.1096/fasebj.10.1.8566530. [DOI] [PubMed] [Google Scholar]
  16. Lee G. J., Pokala N., Vierling E. Structure and in vitro molecular chaperone activity of cytosolic small heat shock proteins from pea. J Biol Chem. 1995 May 5;270(18):10432–10438. doi: 10.1074/jbc.270.18.10432. [DOI] [PubMed] [Google Scholar]
  17. Lee G. J., Roseman A. M., Saibil H. R., Vierling E. A small heat shock protein stably binds heat-denatured model substrates and can maintain a substrate in a folding-competent state. EMBO J. 1997 Feb 3;16(3):659–671. doi: 10.1093/emboj/16.3.659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lee G. J., Vierling E. A small heat shock protein cooperates with heat shock protein 70 systems to reactivate a heat-denatured protein. Plant Physiol. 2000 Jan;122(1):189–198. doi: 10.1104/pp.122.1.189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lindner R. A., Kapur A., Carver J. A. The interaction of the molecular chaperone, alpha-crystallin, with molten globule states of bovine alpha-lactalbumin. J Biol Chem. 1997 Oct 31;272(44):27722–27729. doi: 10.1074/jbc.272.44.27722. [DOI] [PubMed] [Google Scholar]
  20. Lindner R. A., Kapur A., Mariani M., Titmuss S. J., Carver J. A. Structural alterations of alpha-crystallin during its chaperone action. Eur J Biochem. 1998 Nov 15;258(1):170–183. doi: 10.1046/j.1432-1327.1998.2580170.x. [DOI] [PubMed] [Google Scholar]
  21. Martin J., Hartl F. U. Chaperone-assisted protein folding. Curr Opin Struct Biol. 1997 Feb;7(1):41–52. doi: 10.1016/s0959-440x(97)80006-1. [DOI] [PubMed] [Google Scholar]
  22. Rajaraman K., Raman B., Ramakrishna T., Rao C. M. The chaperone-like alpha-crystallin forms a complex only with the aggregation-prone molten globule state of alpha-lactalbumin. Biochem Biophys Res Commun. 1998 Aug 28;249(3):917–921. doi: 10.1006/bbrc.1998.9242. [DOI] [PubMed] [Google Scholar]
  23. Raman B., Ramakrishna T., Rao C. M. Temperature dependent chaperone-like activity of alpha-crystallin. FEBS Lett. 1995 May 29;365(2-3):133–136. doi: 10.1016/0014-5793(95)00440-k. [DOI] [PubMed] [Google Scholar]
  24. Slingsby C., Bateman O. A. Rapid separation of bovine beta-crystallin subunits beta B1, beta B2, beta B3, beta A3 and beta A4. Exp Eye Res. 1990 Jul;51(1):21–26. doi: 10.1016/0014-4835(90)90165-q. [DOI] [PubMed] [Google Scholar]
  25. Treweek T. M., Lindner R. A., Mariani M., Carver J. A. The small heat-shock chaperone protein, alpha-crystallin, does not recognize stable molten globule states of cytosolic proteins. Biochim Biophys Acta. 2000 Aug 31;1481(1):175–188. doi: 10.1016/s0167-4838(00)00109-6. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES