The generation of hydrogen peroxide by the UVA irradiation of human lens proteins.
The water-insoluble proteins from aged human lens are known to contain protein-bound chromophores that act as UVA senisitizers. The irradiation of a sonication-solubilized, water-insoluble fraction from human lenses (55-75 years) with UVA light (1.5 kJ/cm2, gamma > 338nm) caused an oxygen-dependent photolysis of tryptophan, not seen when either alpha-crystallin or lysozyme were irradiated. The suggested requirement for active oxygen species was consistent with a linear increase in hydrogen peroxide formation, which was also observed. A final concentration of 55 microM H2O2 was attained, with no H2O2 being detected in either dark-incubated controls or in irradiated samples of native proteins. The UVA-dependent H2O2 formation was increased 50% by superoxide dismutase (SOD) and abolished by catalase, arguing for the initial generation of superoxide anion. A linear photolysis of histidine and tryptophan was also seen; however, the addition of SOD or SOD and catalase had no effect on the photolytic destruction of either amino acid. Superoxide dismutase increased the oxidation of protein SH groups implicating H2O2, but SOD and catalase caused a decrease in SH oxidation only at later time periods. The direct addition of H2O2 to a water-insoluble sonicate supernatant fraction caused only a slight oxidation of SH groups, but this was increased four- to eight-fold when the protein was denatured in 4.0 M guanidine hydrochloride. Overall, the data suggest a UVA-dependent oxidation of protein SH groups via H2O2 generated within the large protein aggregates of the water-insoluble fraction. These data also provide a mechanism for oxidation of the sulfur-containing amino acids in vivo--a process that is known to accompany the formation of age-onset cataracts.
Linetsky
M
M
Mason Institute of Ophthalmology, University of Missouri, Columbia 65212, USA.
Ortwerth
B J
BJ
D000368:Aged; D003459:Crystallins / Q000737:chemistry / Q000378:metabolism / Q000528:radiation effects*; D005609:Free Radicals; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008875:Middle Aged; D010100:Oxygen / Q000378:metabolism; D010782:Photolysis; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D014364:Tryptophan / Q000378:metabolism; D014466:Ultraviolet Rays
D003459:Crystallins; D005609:Free Radicals; D013438:Sulfhydryl Compounds; D013481:Superoxides; D014364:Tryptophan; D006861:Hydrogen Peroxide; D010100:Oxygen