Nondisulfide polymerization of gamma- and beta-crystallins in the human lens.

Abstract

The water-soluble 43,000-dalton fraction (WS43) of the human lens has been shown to be heterogeneous. It appears to contain, in addition to actin, components related to the crystallins. Immunoblot reactions indicate that this polypeptide fraction is composed of dimers containing (iand y-crystallin components. It has been estimated that 10-30% of this fraction arises by dimerization of y-crystallin. A possible route for the formation of the 43,000-dalton fraction is suggested by the observation that photolysis of y-crystallin with light >295 nm leads to polymer formation, including the 43,000-dalton fraction. The polymerization products react with anti-WS43. The results suggest that photochemical reactions may lead to the accumulation of polymers of some of the crystallins with aging of the human lens. Similar covalently linked polypeptides have previously been shown to be present in the high molecular weight aggregates associated with cataract formation.It is now well established that oxidative insult is fundamentally involved in the cataractous process (1-4). This has been demonstrated by the extensive oxidation of methionine and cysteine that is found during the development of cataract (4). There is also the formation of high molecular weight (HMW) disulfide-linked aggregates that are believed to contribute to the loss of transparency (3). It has been suggested that the aggregation process is initiated at or near the plasma membrane and that a 43,000-dalton extrinsic membrane polypeptide fraction is one of the polypeptides involved in the formation of these aggregates (5).Previous work has shown that the major component containing an atypical protein fluorescence (excitation, 340; 360 nm: emission, 410; 440 nm) was the 43,000-dalton fraction (6). This fluorescence was found to increase with aging and cataract formation and to be present in both soluble and insoluble protein fractions in older lenses (6). Examination of the abundance and distribution of the 43,000-dalton fraction indicated that in very young human lenses, it was only present in the water-insoluble fraction, appearing in the watersoluble fraction at approximately the age of 5 yr and increasing in abundance up to approximately the age of 30 yr where it stabilizes at -8%, a concentration similar to that found in the water-insoluble fraction (7). Studies on the water-insoluble 43,000-dalton fraction indicate that this protein fraction has a similar amino acid composition to the water-soluble 43,000-dalton fraction (WS43) and that these two protein fractions are immunochemically similar (8).What is the origin of this fraction? Lens culture experiments with labeled amino acids demonstrated incorporation into the 43,000-dalton fraction, suggesting polypeptide synthesis (7). The only known polypeptide of this size present in the lens is actin (9, 10). However, the amino acid composition of the 43,000-dalton protein fraction is markedly different from that reported for actin (8). Furthermore, with both Laemmli (11) and Weber-Osborn (12) PAGE s...

Identification of noncollagenous components of calf lens capsule: Evaluation of their adhesion‐promoting activity

“…Laminin and fibronectin eluted from the gel close to the void volume, and these two components were separated from the laterlished the preduminantly noncollagenous character of emerging entactin peak (Kav = 0.30). The P-crystallin, guanidine HCl-extractable proteins and were consistent as anticipated (Roy et al, 1984), appeared as a well with their amino acid composition.…”

Hyperbaric Oxygen in vivo Accelerates the Loss of Cytoskeletal Proteins and MIP26 in Guinea Pig Lens Nucleus

“…The formation of disulfide-crosslinked high molecular weight aggregates, as observed in this study, is a common feature of oxidatively damaged cytoskeletal proteins Graceffa, Adam and Lehman, 1993 ;Schwartz et al, 1987 ;Zaremba et al, 1984). Lens epithelial cells which have been exposed to sulfhydryl oxidants such as H # O # or diamide suffer a severe disruption of cytoskeletal protein organization (Ikebe et al, 1989 ;Prescott et al, 1991), and cytoskeletal components have been reported to be present in the high molecular weight aggregated protein of human senile cataracts (Roy et al, 1984). Two recent studies using oxidative models for cataract in vivo (buthionine sulfoxamine cataract in the mouse and selenite cataract in the rat) have also demonstrated a heightened loss of lens cytoskeletal proteins in the early stages of cataract (Calvin et al, 1992 ;Matsushima et al, 1997).…”

Photooxidation of specific residues in .alpha.-crystallin polypeptides

“…1989; as well as disulfide and non-disulfide covalent cross-linking (Fujimori, 1982;Gossey et al, 1980;Dillon et al, 1985;Roy et al, 1984;Saito & Matsuura, 1985, Schiavon & Veronese, 1986; Borkman et al, 1986). As a result of various therapies and diseases, metabolites such as porphyrins, which are efficient photosensitizers, can reach the eye (Kessel et al, 1987;Fraunfelder, 1982).…”