Heat-induced quaternary transitions in hetero- and homo-polymers of alpha-crystallin.
OBJECTIVE
To compare the effects of heat incubation on the structure and function of native alpha-crystallin, urea denatured/renatured alpha-crystallin, and alphaA and alphaB-crystallin homo-polymers purified from bovine lenses.


METHODS
Each of the alpha-crystallin samples were incubated for 1 h at temperatures ranging from 35 degrees C to 70 degrees C. After heat incubation structural perturbations in each of the samples were studied using non-denaturing gel electrophoresis, transmission electron microscopy (TEM) and far-UV circular dichroism. The chaperone-like activity of each of the heat-treated samples was measured using the DTT induced insulin aggregation assay.


RESULTS
The native alpha-crystallin samples showed secondary structure perturbations, an increase in aggregate size and asymmetry, and an increase in chaperone-like activity after heat incubation above 50 degrees C. The other three sample types showed secondary structure perturbations beginning at lower incubation temperatures, and a progressive decrease in chaperone-like activity with exposure to increasing temperatures. TEM showed all samples formed large asymmetric high molecular weight aggregates after incubation at 65 degrees C.


CONCLUSIONS
The urea denaturation/renaturation of alpha-crystallin has been shown to result in the loss of a small amount of alpha-helix, but to have no effect on chaperone-like activity under standard test conditions. The present results indicate this lost alpha-helix may be responsible for the differential effects of heat incubation on the different forms of alpha-crystallin.
Burgio
M R
MR
Center for Biophysics and Department of Biology, Science Center, Rensselaer Polytechnic Institute, Troy, NY 12180, USA. burgim@rpi.edu
Bennett
P M
PM
Koretz
J F
JF
D000818:Animals; D002417:Cattle; D002942:Circular Dichroism; D003080:Cold Temperature; D003459:Crystallins / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D006358:Hot Temperature; D007908:Lens, Crystalline / Q000737:chemistry*; D018832:Molecular Chaperones / Q000378:metabolism; D011489:Protein Denaturation; D017510:Protein Folding; D020673:Protein Renaturation; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship
D003459:Crystallins; D018832:Molecular Chaperones