title: Metabolism of AF1 (KNEFIRF-NH2) in the nematode, Ascaris suum, by aminopeptidase, endopeptidase and deamidase enzymes.
abstract: We have studied the metabolism and inactivation of AF1 (KNEFIRF-NH2) by membranes prepared from the locomotory muscle of Ascaris suum. FIRF-NH2 and KNEFIRF were identified as three primary degradation products, resulting from the action of an endopeptidase, aminopeptidase and a deamidase, respectively. The endopeptidase resembled mammalian neprilysin (NEP, endopeptidase 24.11) in that the enzyme activity was inhibited by phosphoramidon and thiorphan and that it cleaved AF1 on the amino side of phenylalanine. The aminopeptidase activity was inhibited by amastatin and bestatin but not by puromycin. The deamidation of AF1 was inhibited by phenylmethylsulfonyl fluoride, p-chloromercuricphenylsulfonate and mercuric chloride, indicating that the deamidase enzyme is a serine protease with a requirement for a free thiol group for activity. AF1 (1 microM) induces an increase in tension and an increase in the frequency and amplitude of spontaneous contractions of an A. suum muscle strip. None of the aforementioned AF1 metabolites (2-20 microM) retained biological activity in this bioassay, indicating that the endopeptidase, aminopeptidase and deamidase have the potential to terminate the action of AF1 on locomotory muscle of A. suum.
authors.affiliation: Department of Biology, University of Leeds, UK.
authors.forename: M
authors.initials: M
authors.lastname: Sajid
authors.forename: C
authors.initials: C
authors.lastname: Keating
authors.forename: L
authors.initials: L
authors.lastname: Holden-Dye
authors.forename: I D
authors.initials: ID
authors.lastname: Harrow
authors.forename: R E
authors.initials: RE
authors.lastname: Isaac
mesh_terms: D000581:Amidohydrolases / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D017165:Ascaris suum / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006020:Glycopeptides / Q000494:pharmacology; D015801:Helminth Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D015260:Neprilysin / Q000009:adverse effects / Q000302:isolation & purification / Q000378:metabolism*; D009479:Neuropeptides / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D010455:Peptides; D011480:Protease Inhibitors / Q000494:pharmacology; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D006020:Glycopeptides; D015801:Helminth Proteins; D009124:Muscle Proteins; D009479:Neuropeptides; D010446:Peptide Fragments; D010455:Peptides; D011480:Protease Inhibitors; D015842:Serine Proteinase Inhibitors; C063461:AF1 neuropeptide; C016977:amastatin; D000626:Aminopeptidases; D015260:Neprilysin; D000581:Amidohydrolases; C102793:AF1 deamidase, Ascaris suum; C008890:phosphoramidon