Oxidative modification of the eye lens proteins, the crystallins, is known to cause protein cross-links and aggregates which lead to lens opacification or cataracts. We focus attention here on oxidative damage occurring in crystallins and some "control" proteins upon reaction with the hydroxyl radical (.OH) which, in the lens, is generated by photosensitization or by the Fenton reaction. In the present study, we have synthesized and used the bishydroperoxide I as a "photo-Fenton" reagent, in order to photolytically generate pure .OH, free of other oxyradicals. Our findings are the following: (i) Trp residues are oxidized by .OH to N-formylkynurenine and related compounds, but this in itself does not lead to covalent aggregation of the protein. (ii) Tyr residues react with .OH, but apparently do not produce dihydroxyphenylalanine or bityrosine. Nor do protein cross-links occur as a result. (iii) Oxidation of His residues appears to be obligatory for protein cross-linking. Histidine-free proteins do not form high molecular weight products upon reaction with .OH. Protection of His residues by adduct formation in other proteins inhibits cross-linking. (iv) Lys residues seem to participate in the cross-linking reaction. Protection of the Lys residues by maleylation of the protein inhibits cross-linking. (v) The oxidized protein is more acidic in nature than the parent, and it might have altered conformational features.