Intracellular Localization of Peptide Hydrolases in Wheat (Triticum aestivum L.) Leaves

Abstract

Protoplasts from 8-9 day old wheat (T. aestivum L.) leaves were used to isolate organelles which were examined for their contents of peptide hydrolase enzymes and, in the vacuoles, other acid hydrolases. High yields of intact chloroplasts were obtained using equilibrium density gradient centrifugation and velocity sedimentation centrifugation on sucrose-sorbitol gradients. Aminopeptidase activity was distributed, in approximately equal proportions, between the chloroplasts and cytoplasm. Leucyltyrosine dipeptidase was mainly localized in the cytoplasm, although .apprx. 27% was associated with the chloroplasts. Vacuoles free from cellulysin contamination contained all of the protoplast carboxypeptidase and Hb-degrading activities. The acid hydrolases, phosphodiesterase, acid phosphatase, .alpha.-mannosidase and .beta.-N-acetylglucosaminidase were present in the vacuole to varying degrees, but no .beta.-glucosidase was localized in the vacuole.