title: An improved method for the purification of myrosinase and its physicochemical characterization.
abstract: An improved high yielding procedure for the purification of myrosinase from Sinapis alba L. consisting of concanavalin A affinity chromatography followed by a chromatofocusing step is presented. The purified enzyme was homogeneous as judged by sodium dodecyl sulfate-gel electrophoresis and by analytical ultracentrifugation although the presence of at least three isoenzymes, with pI values from 5.05 to 5.15, was detected by isoelectric focusing. It was found that the enzyme has a molecular weight of 135.1 kg mol-1 and consists of two, possibly identical, subunits of molecular weight 71.7 kg mol-1. The structure of myrosinase was studied by circular dichroism. Contin analysis of the CD data indicates a mixed alpha-helix and beta-sheet conformation for the native protein a with approximately 19% alpha-helix and approximately 35% beta-sheet content. Denaturation with guanidinium chloride was found to be irreversible although the enzyme has excellent storage characteristics in aqueous solution.
authors.affiliation: Institut für Polymere der Eidgenossischen Technischen Hochschule Zürich, ETH-Zentrum, Switzerland.
authors.forename: A
authors.initials: A
authors.lastname: Pessina
authors.forename: R M
authors.initials: RM
authors.lastname: Thomas
authors.forename: S
authors.initials: S
authors.lastname: Palmieri
authors.forename: P L
authors.initials: PL
authors.lastname: Luisi
mesh_terms: D001937:Brassica / Q000201:enzymology*; D055598:Chemical Phenomena; D002621:Chemistry; D002846:Chromatography, Affinity; D006026:Glycoside Hydrolases / Q000302:isolation & purification* / Q000378:metabolism; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D009149:Mustard Plant / Q000201:enzymology*; D010946:Plants, Medicinal
chemical_list: D007527:Isoenzymes; D006026:Glycoside Hydrolases; C022739:thioglucosidase