Processing of enkephalin-containing peptides in isolated bovine adrenal chromaffin granules.

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Abstract
Intact chromaffin granules isolated from bovine adrenal medulla incubated at 37.degree. C for up to 22 h. Processing of enkephalin-containing (EC) peptides in the granules was followed by the change in their size distribution as shown by chromatography on Sephadex G-75 columns. A gradual shift toward lower MW EC peptides was observed during the incubation, indicating processing of the higher MW to lower MW EC peptides. The total amount of [Met]-enkephalin, free and in peptide linkage, remained constant indicating that little or no nonspecific degradation occurred during the experiment. HPLC [high performance liquid chromatography] resolution of the fraction containing the low MW EC peptides showed that free enkephalins as well as [Met]enkephalin-Arg6-Phe7 and [Met]enkephalin-Arg6-Gly7-Leu8 accumulated while [Met]enkephalin-Arg6 and [Met]enkephalin-Lys6 disappeared. The presence of an atypical trypsin activity and the presence of carboxypeptidase B-like activity within the granules is indicated. From the rates of accumulation of the low MW EC peptides and the disappearance of the higher MW EC peptides, a processing rate of 65-70 pmol/g tissue per h was estimated, which calculates to a lifetime of 6-8 days for EC peptides in the granules. Under steady-state conditions this rate of processing appears to be too low to produce significant amounts of free enkephalins from larger EC peptides. This is well in accord with previous observations that relatively small amounts of free enkephalins are found in bovine adrenal medulla.