title: Purification and characterization of acidic endo-polygalacturonase encoded by the PGL1-1 gene from Saccharomyces cerevisiae.
abstract: The PGL1 gene of the yeast Saccharomyces cerevisiae has been shown to encode polygalacturonase. Cloning of the PGL1 open reading frame behind the ADH1 promoter allowed overexpression of polygalacturonase activity in S. cerevisiae. This enzyme was purified to apparent homogeneity from cultures of recombinant S. cerevisiae on synthetic medium using one-step purification by anionic exchange chromatography. The enzyme, named Pgl1P, had an apparent M(r) of 42 kDa as shown by SDS-PAGE. Pgl1P was active from pH 3 to 5.5, with an optimum temperature at 25 degrees C. This enzyme hydrolyzed polygalacturonic acid as an endo-polygalacturonase as demonstrated by independent methods. The purified protein was N-glycosylated. However, the activity remained in the N-deglycosylated form. The N-terminal amino acid sequence was also determined as D-S-C-T-L-T-G-S-S-L.
authors.affiliation: Université de Reims, Faculté des Sciences, Laboratoire de Microbiologie Générale et Moléculaire, Europol'Agro, P.O. Box 1039, 51687, Reims, France.
authors.forename: A
authors.initials: A
authors.lastname: Gainvors
authors.forename: N
authors.initials: N
authors.lastname: Nedjaoum
authors.forename: S
authors.initials: S
authors.lastname: Gognies
authors.forename: M
authors.initials: M
authors.lastname: Muzart
authors.forename: M
authors.initials: M
authors.lastname: Nedjma
authors.forename: A
authors.initials: A
authors.lastname: Belarbi
mesh_terms: D000595:Amino Acid Sequence; D005800:Genes, Fungal; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D011096:Polygalacturonase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development
chemical_list: D011096:Polygalacturonase