Study of the chaperoning mechanism of bovine lens alpha-crystallin, a member of the alpha-small heat shock superfamily.
We have studied the interaction between lysozyme, destabilized by reducing its -S-S- bonds, and bovine eye lens alpha-crystallin, a member of the alpha-small heat shock protein superfamily. We have used gel filtration, photon correlation spectroscopy, and analytical ultracentrifugation to study the binding of lysozyme by alpha-crystallin at 25 degrees C and 37 degrees C. We can conclude that alpha-crystallin chaperones the destabilized protein in a two-step process. First the destabilized proteins are bound by the alpha-crystallin so that nonspecific aggregation of the destabilized protein is prevented. This complex is unstable, and a reorganization and inter-particle exchange of the peptides result in stable and soluble large particles. alpha-Crystallin does not require activation by temperature for the first step of its chaperone activity as it prevents the formation of nonspecific aggregates at 25 degrees C as well as at 37 degrees C. The reorganization of the peptides, however, gives rise to smaller particles at 37 degrees C than at 25 degrees C. Indirect evidence shows that the association of several alpha-crystallin/substrate protein complexes leads to the formation of very large particles. These are responsible for the increase of the light scattering.
Abgar
S
S
Biophysics Research Group, Department of Biochemistry, University of Antwerp, B-2610 Antwerp, Belgium.
Vanhoudt
J
J
Aerts
T
T
Clauwaert
J
J
D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D003459:Crystallins / Q000737:chemistry* / Q000378:metabolism*; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006360:Heat-Shock Proteins / Q000737:chemistry*; D007700:Kinetics; D008027:Light; D015233:Models, Statistical; D009113:Muramidase / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D017785:Photons; D011485:Protein Binding; D021381:Protein Transport; D012542:Scattering, Radiation; D013053:Spectrophotometry; D013696:Temperature; D013997:Time Factors; D014461:Ultracentrifugation
D003459:Crystallins; D006360:Heat-Shock Proteins; D010455:Peptides; D009113:Muramidase; D004229:Dithiothreitol