The localization of lysosomal enzymes in chromaffin tissue

Abstract

An homogenate of bovine adrenal medulla contains significant amounts of 6 acid hydrolases: acid ribonuclease, acid deoxyribonuclease, cathepsin, acid phosphatase, [beta]-glucuronidase and arylsulphatase. Most of the activity of each enzyme could be sedimented in the large-granule fraction at 242,000 g-min. Differential centrifugation indicated the presence of 3 populations of particles, which sedimented at slightly different rates; these are, in order of decreasing sedimentation rate, mitochondria, particles containing the acid hydrolases, and chromaffin granules. The 3 types of particle could be separated by ultracentrifuging the large-granule fraction in a sucrose density gradient. Most of the activity of each hydrolase was recovered in a layer intermediate between those formed by mitochondria and chromaffin granules. The large-granule fraction therefore contains particles which are defined by their enzyme content as lysosomes. Highly purified chromaffin granules, containing less than 5% of the activity of each acid hydrolage, were obtained from the gradient.