Ion-exchange chromatographic assay of peptidases acting on the C-terminal hexapeptide sequence of substance P.
A rapid and sensitive assay for peptidases acting on the C-terminal hexapeptide sequence of the neuropeptide substance P is described. The radiolabelled substrate, N alpha-[125I]desaminotyrosyl-substance P (6-11) is easily prepared by coupling commercially available radioiodinated Bolton-Hunter reagent with substance P (6-11). Peptidase activity is determined by quantitative separation of the degradation products from the intact substrate on small QAE-Sephadex columns. The assay has been used to measure degradation of the substrate by rat parotid and diencephalon slices. The peptidase activity in the latter system was inhibited by substance P and substance P fragments and was sensitive to metal chelators and thiol reagents.
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D004027:Diencephalon / Q000032:analysis; D006168:Guinea Pigs; D007700:Kinetics; D009130:Muscle, Smooth / Q000032:analysis; D009419:Nerve Tissue Proteins / Q000032:analysis; D010306:Parotid Gland / Q000032:analysis; D010443:Peptide Chain Termination, Translational; D010447:Peptide Hydrolases / Q000032:analysis* / Q000378:metabolism; D013373:Substance P / Q000378:metabolism*
D009419:Nerve Tissue Proteins; D013373:Substance P; D010447:Peptide Hydrolases