Effect of pleiotropic molecules on alpha-crystallin aggregation

Crystallins are the predominant proteins within the eye lens. In mammals, the crystallins family is composed of three classes denoted a, b and c. α-Crystallin, the principal lens protein, belongs to the heat shock proteins family; it acts as a molecular chaperone, being thus crucial to prevent the aggregation and/or precipitation of other partially denatured proteins, itself included due to a self-chaperone property. In its native state a-crystallin consists of two relatively homologous subunits αA and αB, each of about 20 kDa and forms large, heterogeneous, water-soluble low-molecular weight (LMW) aggregates (MW about 800 kDa). Under certain conditions, included normal aging and/or various stresses, may progressively became part of insoluble high-molecular weight (HMW) systems aggregates (more than 1000 kDa). These systems are supposed to play a relevant role in eye lens opacification and vision impairment. To stabilize protein folding, inhibit and prevent protein aggregation and modification, several small molecules have been studied. Among these, trehalose, a nonreducing disaccharide composed of two glucose molecules, and carnosine, a natural dipeptide (ß-alanyl-histidine) with potential protective and therapeutic significance in many diseases, have been shown protective effects. We report the pleitropic effects of trehalose and carnosine, on α crystallin aggregation. The α-crystallin stress tolerance, chaperone activity and thermal stability is studied by calorimetric (DSC) microscopic (AFM) and fluorescence analyses measurements (ThT). Under in vitro denaturing conditions the presence of Trehalose was able to stabilize the α-crystallin native structure, to inhibit α-crystallin aggregation, and to disaggregate preformed LMW systems not affecting the chaperone activity. L-carnosine, has been proposed as potent agents for ophthalmic therapies of senile cataracts and diabetic ocular complications. We observed inhibitory effect induced by the peptide on crystallin fibrillation and an almost complete restoration of the chaperone activity lost after denaturant and/or heat stress. Moreover, we found a disassembling effect of carnosine on crystallin amyloid fibrils, as well as the ability of carnosine to restore most of the lens transparency in organ-cultured rat lenses exposed to denaturing conditions.