Specificity of neuropeptide degradation by two calcium-activated neutral proteases from human skeletal muscle.
Two calcium-activated neutral proteases (CAPI & II) were purified from human skeletal muscle by anion exchange, gel filtration and affinity (antipain-Sepharose and Blue Ultrogel A4R) chromatography. The enzymes were homogenous as judged by polyacrylamide gel electrophoresis, and have similar properties with the exception of the Ca2+ concentration required for optimum activity (CAP I = 0.1 mM; CAP II = 1 mM). Both enzymes hydrolysed a wide variety of neuropeptides. In six cases, the products were separated and identified by hplc and amino acid analysis. Neurotensin was hydrolysed at Tyr3-Glu4; dynorphin1-13 at Arg8-Arg9; LH-RH at Gly6-Leu7; CCK-8 at Phe8-NH2, substance-P at Met10-NH2; somatostatin at Thr10-Phe11. Although differences in the rates of neuropeptide degradation were noted for the two CAP's the specificity was the same for these six peptides. It is suggested that conformational requirements may be more important than side chains adjacent to the cleavage site in directing the specificity of CAP.
J R
JR
McDermott
D
D
Mantle
J A
JA
Biggins
A M
AM
Kidd
K
K
Davison
B
B
Lauffart
R J
RJ
Pennington
D000596:Amino Acids / Q000032:analysis; D002154:Calpain; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004399:Dynorphins / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D007987:Gonadotropin-Releasing Hormone / Q000032:analysis; D006801:Humans; D009132:Muscles / Q000201:enzymology*; D009419:Nerve Tissue Proteins / Q000032:analysis*; D009496:Neurotensin / Q000032:analysis; D012844:Sincalide / Q000032:analysis; D013373:Substance P / Q000032:analysis
D000596:Amino Acids; D009419:Nerve Tissue Proteins; D013373:Substance P; D007987:Gonadotropin-Releasing Hormone; D009496:Neurotensin; D004399:Dynorphins; D010450:Endopeptidases; D002154:Calpain; D012844:Sincalide