Comparative analysis of prosomes and multicatalytic proteinases from rabbit erythrocytes

Abstract

Prosomes and multicatalytic proteinases were purified from rabbit erythrocyte lysates and were analysed to determine their relationship. During purification by sucrose density gradient centrifugation using low salt buffer, they sedimented at 20–26S. Upon further purification, using high salt buffer, prosomes were recovered as 20S complexes as determined by their characteristic polypeptide pattern. Interestingly, both the 26S and 20S components had protease activity. Therefore, in order to determine their relationship with the multicatalytic proteinases, which are reported to contain a similar set of polypeptides, highly pure prosomes and the multicatalytic proteinases were analysed. Both 20S prosomes and multicatalytic proteinases showed protease activity and also had identical protein subunits of molecular weight ranging from 21 kDa to 35 kDa. Among these, at least two were immunologically identical as determined by Western blot using two monoclonal antibodies prepared against duck prosomes. Furthermore, protease activities of both components were inhibited almost to the same extent by an endogenous inhibitor specific for high molecular weight proteases and calpain. These results thus establish that the 20S prosomes and multicatalytic proteinases are identical, and suggest further that proteolytic activity could be the principal function of prosomes.