title: Purification and characterization of a glutathione S-transferase from Mucor mucedo.
abstract: An intracellular glutathione transferase was purified to homogenity from the fungus, Mucor mucedo, using DEAE-cellulose ion-exchange and glutathione affinity chromatography. Gel filtration chromatography and SDS-PAGE revealed that the purified GST is a homodimer with approximate native and subunit molecular mass of 53 kDa and 23.4 kDa, respectively. The enzyme has a pI value of 4.8, a pH optimum at pH 8.0 and apparent activation energy (Ea) of 1.42 kcal mol(-1). The purified GST acts readily on CDNB with almost negligible peroxidase activity and the activity was inhibited by Cibacron Blue (IC50 0.252 microM) and hematin (IC50 3.55 microM). M. mucedo GST displayed a non-Michaelian behavior. At low (0.1-0.3 mM) and high (0.3-2 mM) substrate concentration, Km (GSH) was calculated to be 0.179 and 0.65 mM, whereas Km(CDNB) was 0.531 and 11 mM and k(cat) was 39.8 and 552 s(-1), respectively. The enzyme showed apparent pKa values of 6-6.5 and 8.0.
authors.affiliation: Molecular Biology Department, National Research Centre, Cairo, Egypt. hamed_rr@hotmail.com
authors.forename: Ragaa R
authors.initials: RR
authors.lastname: Hamed
authors.forename: Mohamed R
authors.initials: MR
authors.lastname: Abu-Shady
authors.forename: Fawkia M
authors.initials: FM
authors.lastname: El-Beih
authors.forename: Abdel-Monem A
authors.initials: AM
authors.lastname: Abdalla
authors.forename: Ola M
authors.initials: OM
authors.lastname: Afifi
mesh_terms: D002850:Chromatography, Gel; D005982:Glutathione Transferase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009089:Mucor / Q000201:enzymology* / Q000254:growth & development
chemical_list: D005982:Glutathione Transferase