Age-related modifications in rat cardiac phosphoglycerate kinase. Rejuvenation of the old enzyme by unfolding-refolding

Abstract

The occurrence of age-related modifications in functional and structural properties of several enzymes has been documented; however, the molecular basis of this phenomenon is still mostly unexplained. In the present work a comparative study of phosphoglycerate kinase preparations isolated from hearts of young and old rats was undertaken. Marked age-related effects wer revealed in the heat-inactivation kinetics of the enzyme, similar to the ones previously found in purified muscle phosphoglycerate kinase. In view of the previously reported failure of immunotitration to distinguish between phosphoglycerate kinase forms in crude heart extracts from young and old rats, it appears likely that the modifications in old rat heart phosphoglycerate kinase are in a domain which is not involved in antibody binding, and may be localized in the interior of the enzyme. These age-related modifications were completely relieved by extensive unfolding of the enzyme in 2 M guanidine hydrochloride, followed by enzyme reactivation upon dilution of the denaturant. The refolding products of young and old enzymes displayed identical heat-inactivation kinetics as native young phosphoglycerate kinase. It is concluded that the age-related alterations in rat cardiac phosphoglycerate kinase, like those found in the muscle enzyme, are purely conformational and hence develop postsynthetically.