Prohormone thiol protease and enkephalin precursor processing: cleavage at dibasic and monobasic sites.
Production of active enkephalin peptides requires proteolytic processing of proenkephalin at dibasic Lys-Arg, Arg-Arg, and Lys-Lys sites, as well as cleavage at a monobasic arginine site. A novel "prohormone thiol protease" (PTP) has been demonstrated to be involved in enkephalin precursor processing. To find if PTP is capable of cleaving all the putative cleavage sites needed for proenkephalin processing, its ability to cleave the dibasic and the monobasic sites within the enkephalin-containing peptides, peptide E and BAM-22P (bovine adrenal medulla docosapeptide), was examined in this study. Cleavage products were separated by HPLC and subjected to microsequencing to determine their identity. PTP cleaved BAM-22P at the Lys-Arg site between the two basic residues. The Arg-Arg site of both peptide E and BAM-22P was cleaved at the NH2-terminal side of the paired basic residues to generate [Met]-enkephalin. Furthermore, the monobasic arginine site was cleaved at its NH2-terminal side by PTP. These findings, together with previous results showing PTP cleavage at the Lys-Lys site of peptide F, demonstrate that PTP possesses the necessary specificity for all the dibasic and monobasic cleavage sites required for proenkephalin processing. In addition, the unique specificity of PTP for cleavage at the NH2-terminal side of arginine at dibasic or monobasic sites distinguishes it from many other putative prohormone processing enzymes, providing further evidence that PTP appears to be a novel prohormone processing enzyme.
Krieger
T J
TJ
Department of Biochemistry, Uniformed Services University of the Health Sciences, Bethesda, Maryland 20814.
Mende-Mueller
L
L
Hook
V Y
VY
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004744:Enkephalin, Methionine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D004745:Enkephalins / Q000378:metabolism*; D008969:Molecular Sequence Data; D010452:Peptide Biosynthesis; D010455:Peptides / Q000737:chemistry; D011498:Protein Precursors / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D011499:Protein Processing, Post-Translational
D004745:Enkephalins; D010455:Peptides; D011498:Protein Precursors; C029992:proenkephalin; D004744:Enkephalin, Methionine; C028837:BAM 22P; C039933:peptide E (adrenal medulla); D003546:Cysteine Endopeptidases