Pantethine inhibits the formation of high-Tc protein aggregates in gamma B crystallin solutions.

PURPOSE: Solutions of the bovine lens protein gamma B (or gamma II) crystallin at neutral pH in the absence of reducing agents, undergo a slow, partial conversion to a new protein species, gamma IIH. This species is an aggregate composed of an intermolecular, disulfide-crosslinked dimer (approximately equal to 32% of total protein by weight) and loosely associated dimers (approximately equal to 66%). gamma IIH has a phase separation temperature (Tph), at least 40 degrees C higher than that of native gamma II crystallin at any given protein concentration. In this paper we demonstrate that pantethine, a derivative of coenzyme A, inhibits the formation of gamma IIH. METHODS: gamma II crystallin solutions were incubated at pH 7.1 and room temperature with increasing amounts of pantethine. The Tph of the solutions was monitored as a function of incubation time. Corresponding to each Tph measurement, aliquots of each solution were analyzed by cation-exchange HPLC to determine the amount of gamma IIH formed. RESULTS: Incubation of gamma II crystallin with increasing amounts of pantethine lowers Tph and suppresses the formation of gamma IIH. With pantethine to protein mole ratios of 0.66, 1 and 2, the Tph of gamma II crystallin is lowered from 8 degrees C in the native protein, to 2 degrees C, -3 degrees C respectively, at a protein concentration of approximately equal to 200 mg/ml. The amount of gamma IIH accumulated decreases from approximately 25% in the native protein to 10%, 1% and 0% respectively in these pantethine-treated protein solutions. For complete suppression of the rise in Tph and inhibition of gamma IIH formation, a 2:1 mole ratio of pantethine to protein is required. CONCLUSIONS: We suggest that pantethine reacts with two cysteine residues of gamma IIH crystallin by forming a mixed disulfide, and effectively suppress protein aggregation and lowers Tph. This is due to the strong polar character of pantethine which reduces the net attractive interactions between the protein molecules.