title: Aged and non-aged pyrenebutyl-containing organophosphoryl conjugates of chymotrypsin. Preparation and comparison by 31P-NMR spectroscopy.
abstract: Homologous pairs of non-aged and aged pyrene-containing phosphoryl conjugates of chymotrypsin were prepared in order to characterize by NMR and optical spectroscopy putative differences in the conformation of non-aged and aged organophosphoryl conjugates of serine hydrolases. Pyrenebutyl-O-P(O)(OC2H5)F and pyrenebutyl-O-P(O)(OC2H5)Cl were used to obtain the non-aged form pyrenebutyl-O-P(O)(OC2H5)-Cht, whereas pyrenebutyl-O-P(O)Cl2, pyrenebutyl-O-P(O)(p-nitrophenoxy)Cl, and pyrenebutyl-O-P(O)(p-nitrophenoxy)2 were used to produce the aged conjugate pyrenebutyl-O-P(O)(O )-Cht. These ligands bind covalently to the active site of serine hydrolases. The absorption spectra of both the non-aged and aged conjugates fitted approximately a 1:1 stoichiometry of bound organophosphate and enzyme in the non-aged and aged conjugates. Pyrenebutyl-O-P(O)(OC2H5)-Cht could be reactivated by pyridine-3-aldoxime methiodide, whereas no reactivation was observed for the similarly treated pyrenebutyl-O-P(O)(O-)-Cht. The 31P-NMR and reactivation data taken together strongly support the hypothesis that the aged form of the OP-Cht conjugate contains a P--O- bond. These results provide a partial interpretation for the known resistance of the aged conjugates of serine hydrolases to reactivation.
authors.affiliation: Israel Institute for Biological Research, Ness-Ziona.
authors.forename: J
authors.initials: J
authors.lastname: Grunwald
authors.forename: Y
authors.initials: Y
authors.lastname: Segall
authors.forename: E
authors.initials: E
authors.lastname: Shirin
authors.forename: D
authors.initials: D
authors.lastname: Waysbort
authors.forename: N
authors.initials: N
authors.lastname: Steinberg
authors.forename: I
authors.initials: I
authors.lastname: Silman
authors.forename: Y
authors.initials: Y
authors.lastname: Ashani
mesh_terms: D002918:Chymotrypsin / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000138:chemical synthesis; D003640:Dealkylation; D004789:Enzyme Activation; D005453:Fluorescence; D009682:Magnetic Resonance Spectroscopy; D009943:Organophosphorus Compounds / Q000032:analysis / Q000138:chemical synthesis* / Q000494:pharmacology; D011487:Protein Conformation; D011721:Pyrenes / Q000032:analysis
chemical_list: D009943:Organophosphorus Compounds; D011721:Pyrenes; D002918:Chymotrypsin