The damaging effect of UV-C irradiation on lens alpha-crystallin.
OBJECTIVE
To evaluate the effect of UV-C irradiation on the structural properties of alpha-crystallin and its chaperone activity.


METHODS
alpha- and betaL-crystallins were isolated from bovine lenses using gel chromatography. The purified alpha-crystallin was subjected to UV-C irradiation (254 nm; 1, 2, 5, 10, 20, 50 J/cm2). We measured the tryptophan fluorescence, circular dichroism (CD) spectroscopy in the far UV, and the chaperone activity of both irradiated and non-irradiated alpha-crystallin.


RESULTS
The tryptophan fluorescence of alpha-crystallin decreased, whereas the N-formylkynurenine fluorescence increased markedly with increasing doses of UV-C irradiation. Both the oxidation of Met1 and the racemization of Asp151 of alphaA-crystallin increased at a dose of 1-2 J/cm2 and then gradually decreased. The CD spectrum showed that the secondary structure of alpha-crystallin altered with increasing radiation dose, and almost all of the beta-sheet structure was lost at doses above 50 J/cm2. The chaperone activity of alpha-crystallin irradiated with doses under 5 J/cm2 remained intact. However, it was reduced to only 40% after irradiation at 10 J/cm2.


CONCLUSIONS
Our study suggests that photo-oxidation of tryptophan residues in alpha-crystallin may be one of the events that affects the three-dimensional packing array and chaperone activity of this lens protein.
Fujii
Noriko
N
Research Reactor Institute, Kyoto University, Sennan, Osaka, Japan. nfujii@hl.rri.kyoto-u.ac.jp
Uchida
Hiroki
H
Saito
Takeshi
T
D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D002942:Circular Dichroism; D007737:Kynurenine / Q000031:analogs & derivatives* / Q000378:metabolism; D007908:Lens, Crystalline / Q000737:chemistry / Q000378:metabolism / Q000528:radiation effects*; D018832:Molecular Chaperones / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014364:Tryptophan / Q000378:metabolism; D014466:Ultraviolet Rays; D038201:alpha-Crystallins / Q000302:isolation & purification / Q000378:metabolism / Q000528:radiation effects*
D018832:Molecular Chaperones; D038201:alpha-Crystallins; C007772:N'-formylkynurenine; D007737:Kynurenine; D014364:Tryptophan