title: Steroid adduct formation with lens crystallins.
abstract: BACKGROUND: Development of steroid cataract is a likely outcome following prolonged exposure to glucocorticoids. It has been suggested that formation of steroid-protein adducts is a key event in this lens opacification. In order to explore this possibility, we have monitored the reaction of bovine lens proteins with glucocorticoids and examined the effects of adduct formation on their structures. METHODS: Bovine lens proteins were incubated with high (10(-4) M) and low (10(-8) M) concentrations of dexamethasone or prednisolone for up to 56 days at 37 degrees Celsius. Changes in molecular size and solubility of the crystallins and their polypeptide subunits were examined using gel permeation chromatography and SDS gel electrophoresis. Conformational changes were assessed with the aid of tryptophan fluorescence spectroscopy and oxidation was monitored by measuring protein sulphydryl content. RESULTS: Covalent incorporation of glucocorticoids was observed for all crystallins with relative reactivities for alpha-: beta-: gamma-crystallin of 20: 5: 1. The maximum incorporated was one steroid molecule per 40 to 50 subunits of alpha-crystallin. The proportions and sizes of the soluble crystallins and their subunits were unchanged. Protein sulphydryl contents decreased by eight to 10 per cent more than controls but no intermolecular disulphide bonds were detected. There were no alterations in tryptophan microenvironments. CONCLUSIONS: Steroids form adducts with lens proteins, in particular alpha-crystallin, but it appears unlikely that this reaction is responsible for steroid cataract formation.
authors.affiliation: National Vision Research Institute of Australia, 386 Cardigan Street, Carlton, Victoria, 3053, Australia.
authors.forename: Andrew I
authors.initials: AI
authors.lastname: Jobling
authors.forename: Robert C
authors.initials: RC
authors.lastname: Augusteyn