title: Chemical modification of alpha crystallin.
abstract: Calf lens alpha-crystallin was isolated and the lysine residues were extensively modified with a variety of chemical agents. The effect of these modifications on elastase inhibitor activity, apparent molecular size, antibody reactivity and solubility were determined. The addition of either a methyl group or a threose residue did not alter the charge on the lysine residues and had little or no effect on either inhibitor activity or apparent molecular size. The introduction of a negative charge by either carboxymethylation or citraconylation caused a marked decrease in size and an almost complete loss of inhibitor activity. The introduction of a hydrophobic residue by reaction with either a trinitrobenzenesulfonic acid or Bolton Hunter reagent caused a slight increase in size, but a 70% increase in elastase inhibitor activity. Reaction with fluorescamine resulted in the dissociation of alpha-crystallin in a 200-kDa species, yet caused a two to four-fold increase in elastase inhibitor activity, which was similar to the activity of the water-insoluble fraction isolated from aged human lens and cataract. Several of these modified alpha-crystallins were compared for reactivity with a polyclonal alpha-crystallin antiserum using a quantitative slot blot assay. Charge neutral modifications resulted in a two to three-fold loss of antibody recognition, whereas the other preparations showed an almost complete loss of antigenic activity. None of the modifications caused the alpha-crystallin to precipitate at higher salt concentrations (0.3 M) with the exception of threose which caused a 30% decrease in soluble protein.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Mason Institute of Ophthalmology, University of Missouri, Columbia 65212.
authors.forename: B J
authors.initials: BJ
authors.lastname: Ortwerth
authors.forename: P R
authors.initials: PR
authors.lastname: Olesen
authors.forename: K K
authors.initials: KK
authors.lastname: Sharma
authors.forename: M
authors.initials: M
authors.lastname: Prabhakaram
mesh_terms: D000818:Animals; D001666:Binding Sites, Antibody / Q000187:drug effects; D002417:Cattle; D003459:Crystallins / Q000187:drug effects*; D008970:Molecular Weight; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors; D012995:Solubility; D013329:Structure-Activity Relationship
chemical_list: D003459:Crystallins; D010196:Pancreatic Elastase