title: Effect of dicarbonyl-induced browning on alpha-crystallin chaperone-like activity: physiological significance and caveats of in vitro aggregation assays.
abstract: Alpha-crystallin is a member of the small heat-shock protein family and functions like a molecular chaperone, and may thus help in maintaining the transparency of the eye lens by protecting the lens proteins from various stress conditions. Non-enzymic glycation of long-lived proteins has been implicated in several age- and diabetes-related complications, including cataract. Dicarbonyl compounds such as methylglyoxal and glyoxal have been identified as the predominant source for the formation of advanced glycation end-products in various tissues including the lens. We have investigated the effect of non-enzymic browning of alpha-crystallin by reactive dicarbonyls on its molecular chaperone-like function. Non-enzymic browning of bovine alpha-crystallin in vitro caused, along with altered secondary and tertiary structures, cross-linking and high-molecular-mass aggregation. Notwithstanding these structural changes, methylglyoxal- and glyoxal-modified alpha-crystallin showed enhanced anti-aggregation activity in various in vitro aggregation assays. Paradoxically, increased chaperone-like activity of modified alpha-crystallin was not associated with increased surface hydrophobicity and rather showed less 8-anilinonaphthalene-l-sulphonic acid binding. In contrast, the chaperone-like function of modified alpha-crystallin was found to be reduced in assays that monitor the prevention of enzyme inactivation by UV-B and heat. Moreover, incubation of bovine lens with methylglyoxal in organ culture resulted in cataract formation with accumulation of advanced glycation end-products and recovery of alpha-crystallin in high proportions in the insoluble fraction. Furthermore, soluble alpha-crystallin from methylglyoxal-treated lenses showed decreased chaperone-like activity. Thus, in addition to describing the effects of methylglyoxal and glyoxal on structure and chaperone-like activity, our studies also bring out an important caveat of aggregation assays in the context of the chaperone function of alpha-crystallin.
authors.affiliation: National Institute of Nutrition, Hyderabad 500 007, India.
authors.forename: M Satish
authors.initials: MS
authors.lastname: Kumar
authors.forename: P Yadagiri
authors.initials: PY
authors.lastname: Reddy
authors.forename: P Anil
authors.initials: PA
authors.lastname: Kumar
authors.forename: Ira
authors.initials: I
authors.lastname: Surolia
authors.forename: G Bhanuprakash
authors.initials: GB
authors.lastname: Reddy
mesh_terms: D000818:Animals; D002417:Cattle; D017127:Glycation End Products, Advanced / Q000096:biosynthesis / Q000737:chemistry; D006037:Glyoxal / Q000494:pharmacology*; D057927:Hydrophobic and Hydrophilic Interactions; D007908:Lens, Crystalline / Q000033:anatomy & histology / Q000187:drug effects; D015416:Maillard Reaction; D009924:Organ Culture Techniques; D011487:Protein Conformation; D021122:Protein Subunits / Q000737:chemistry; D011765:Pyruvaldehyde / Q000494:pharmacology; D038201:alpha-Crystallins / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*
chemical_list: D017127:Glycation End Products, Advanced; D021122:Protein Subunits; D038201:alpha-Crystallins; D006037:Glyoxal; D011765:Pyruvaldehyde