bam-12p and adrenorphin

An enzymatic activity has been identified which is capable of generating a product chromatographically identical with adrenorphin from the model substrate BAM12P. This enzyme was purified by gel filtration and ion-exchange chromatography and characterised as having a molecular mass between 30 and 45 kDa and an acidic pI. The enzyme is active at the acid pH expected in the secretory vesicle interior and is inhibited by EDTA, suggesting that it is a metalloprotease. This activity could not be mimicked by incubation with lysosomal fractions and it meets the criteria to be considered as a possible prohormone processing enzyme.

Topics: Adrenal Medulla; Animals; Cell Fractionation; Centrifugation, Density Gradient; Chromaffin Granules; Chromatography, Gel; Chromatography, High Pressure Liquid; Chromatography, Ion Exchange; Dithiothreitol; Edetic Acid; Endopeptidases; Enkephalin, Methionine; Metalloendopeptidases; Molecular Weight; Protease Inhibitors; Protein Precursors; Sheep

Met5-enkephalin- (Met-ENK), Leu5-enkephalin-, Met5-enkephalin-Arg6-Phe7-, metorphamide- and BAM 22P-like peptides could be detected in the lumbar spinal cord of the chicken by immunocytochemistry and/or high performance liquid chromatography. However, a peptide identical to Met5-enkephalin-Arg6-Gly7-Leu8 could not be detected in the lumbar spinal cord of the chicken using an antiserum that was capable of detecting the octapeptide in mammalian tissues. Nerve fiber- and terminal-like processes containing proenkephalin-derived peptides were concentrated in the superficial laminae of the dorsal horn and along the midline rostral to the central canal. A lesser concentration of processes containing proenkephalin-derived peptides occurred in the medial and lateral motor columns of the ventral horn. The level of total radioimmunoassayable Met-ENK in the lumbar spinal cord of the chicken embryo increased more than 1000-fold between day 4.5 and day 18. A schedule of curare administration that had previously been shown to prevent naturally occurring somatic motoneuron death in the chicken lumbar spinal cord resulted in a two-fold increase in total radioimmunoassayable Met-ENK in the lumbar spinal cord.

Topics: Animals; Chemical Phenomena; Chemistry; Chickens; Enkephalin, Leucine; Enkephalin, Methionine; Enkephalins; Immunoenzyme Techniques; Protein Precursors; Spinal Cord