title: Production, purification, and characterization of a β-glucosidase of Penicillium funiculosum NCL1.
abstract: Penicillium funiculosum NCL1, a filamentous fungus, produced significantly higher levels of β-glucosidase. The effect of initial pH, incubation temperature, and different carbon sources on extracellular β-glucosidase production was studied in submerged fermentation. At 30 °C with initial pH 5.0, enzyme production was increased by 48-fold upon induction with paper mill waste, as compared to commercial cellulose powder. In zymogram analysis, four isoforms of β-glucosidases were observed with wheat bran whereas a minimum of one isoform was observed with other carbon sources. A major β-glucosidase (Bgl3A) with the apparent molecular weight of ~120 kDa, induced by paper mill waste, was purified 19-fold to homogeneity, with a specific activity of 1,796 U/mg. Bgl3A was a monomeric glycoprotein with 29% of neutral carbohydrate content. It showed optimum activity at pH 4.0 and 5.0, optimum temperature at 60 °C, and exhibited a half-life of 1 h at 60 °C. K(m) of Bgl3A was found to be 0.057 mM with p-nitrophenyl β-D-glucoside and V(max) was 1,920 U/mg. The purified enzyme exhibited glucose tolerance with a K(i) of 1.5 mM. Bgl3A readily hydrolyzed glucosides with β-linkage. Bgl3A activity was enhanced (156%) by Zn²⁺ and was not affected by other metal cations and reagents. The supplementation of Bgl3A (5 U/mg) with Trichoderma reesei cellulase complex (5 FPU/mg) resulted in about 70% of enhanced glucose production, which emphasizes the industrial importance of Bgl3A.
authors.affiliation: Department of Genetics, Centre for Excellence in Genomic Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625021, India.
authors.forename: Gurusamy
authors.initials: G
authors.lastname: Ramani
authors.forename: Balasubramanian
authors.initials: B
authors.lastname: Meera
authors.forename: Chinnathambi
authors.initials: C
authors.lastname: Vanitha
authors.forename: Mala
authors.initials: M
authors.lastname: Rao
authors.forename: Paramasamy
authors.initials: P
authors.lastname: Gunasekaran
mesh_terms: D055162:Biocatalysis; D002244:Carbon / Q000378:metabolism; D002480:Cellulase / Q000378:metabolism; D002482:Cellulose / Q000378:metabolism; D005285:Fermentation; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007527:Isoenzymes / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D010407:Penicillium / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature; D014242:Trichoderma / Q000201:enzymology; D001617:beta-Glucosidase / Q000096:biosynthesis* / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D007527:Isoenzymes; D002244:Carbon; D002482:Cellulose; D001617:beta-Glucosidase; D002480:Cellulase