Tissue kallikrein processes small proenkephalin peptides.
Tissue kallikrein may play a role in processing precursor polypeptide hormones. We investigated whether hydrolysis of natural enkephalin precursors, peptide F and bovine adrenal medulla docosapeptide (BAM-22P), by hog pancreatic kallikrein is consistent with this concept. Incubation of peptide F with this tissue kallikrein resulted in the release of Met5-enkephalin and Met5-Lys6-enkephalin. Met5-Lys6-enkephalin was the main peptide released, indicating that the major cleavage site was between two lysine residues. At 37 degrees C and pH 8.5, the KM values for formation of Met5-enkephalin and Met5-Lys6-enkephalin were 129 and 191 microM, respectively. Corresponding kcat values were 0.001 and 0.03 s-1 and kcat/KM ratios were 8 and 1.6.10(2) M-1.s-1, respectively. Cleavage of peptide F at acidic pH (5.5) was negligible. When BAM-22P was used as a substrate, Met5-Arg6-enkephalin was released, thus indicating cleavage between two arginine residues. At pH 8.5, KM was 64 microM, kcat was 4.5 s-1, and the kcat/KM ratio was 7.10(4) M-1.s-1. At 5.5, the pH of the secretory granules, KM, kcat and kcat/KM were 184 microM, 1.9 s-1 and 10(4) M-1.s-1, respectively. It is unlikely that peptide F could be a substrate for kallikrein in vivo; however, tissue kallikrein could aid in processing proenkephalin precursors such as BAM-22P by cleaving Arg-Arg peptide bonds.
Mindroiu
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Hypertension Research Division, Henry Ford Hospital, Detroit, MI 48202.
Carretero
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Walz
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Scicli
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D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004744:Enkephalin, Methionine / Q000031:analogs & derivatives / Q000378:metabolism; D004745:Enkephalins / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007610:Kallikreins / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology; D011498:Protein Precursors / Q000378:metabolism*; D013552:Swine
D000596:Amino Acids; D004745:Enkephalins; D011498:Protein Precursors; C029992:proenkephalin; D004744:Enkephalin, Methionine; C036452:peptide F; C028837:BAM 22P; D007610:Kallikreins