title: A novel β-glucosidase from Humicola insolens with high potential for untreated waste paper conversion to sugars.
abstract: Humicola insolens produced a new β-glucosidase (BglHi2) under solid-state fermentation. The purified enzyme showed apparent molecular masses of 116 kDa (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and 404 kDa (gel-filtration), suggesting that it is a homotetramer. Mass spectrometry analysis showed amino acid sequence similarity with a β-glucosidase from Chaetomium thermophilum. Optima of pH and temperature were 5.0 and 65 °C, respectively, and the enzyme was stable for 60 min at 50 °C, maintaining 71 % residual activity after 60 min at 55 °C. BglHi2 hydrolyzed p-nitrophenyl-β-D-glucopyranoside and cellobiose. Cellobiose hydrolysis occurred with high apparent affinity (K M = 0.24 ± 0.01 mmol L(-1)) and catalytic efficiency (k cat/K M = 1,304.92 ± 53.32 L mmol(-1) s(-1)). The activity was insensitive to Fe(+3), Cr(+2), Mn(+2), Co(+2), and Ni(2+), and 50-60 % residual activities were retained in the presence of Pb(2+), Hg(2+), and Cu(2+). Mixtures of pure BglHi2 or H. insolens crude extract (CE) with crude extracts from Trichoderma reesei fully hydrolyzed Whatman no. 1 paper. Mixtures of H. insolens CE with T. reesei CE or Celluclast 1.5 L fully hydrolyzed untreated printed office paper, napkin, and magazine papers after 24-48 h, and untreated cardboard was hydrolyzed by a H. insolens CE/T. reesei CE mixture with 100 % glucose yield. Data revealed the good potential of BglHi2 for the hydrolysis of waste papers, promising feedstocks for cellulosic ethanol production.
authors.affiliation: Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, 14040-901, SP, Brazil.
authors.forename: Luana Parras
authors.initials: LP
authors.lastname: Meleiro
authors.forename: Ana Lucia Ribeiro Latorre
authors.initials: AL
authors.lastname: Zimbardi
authors.forename: Flavio Henrique Moreira
authors.initials: FH
authors.lastname: Souza
authors.forename: Douglas Chodi
authors.initials: DC
authors.lastname: Masui
authors.forename: Tony Marcio
authors.initials: TM
authors.lastname: Silva
authors.forename: João Atilio
authors.initials: JA
authors.lastname: Jorge
authors.forename: Rosa Prazeres Melo
authors.initials: RP
authors.lastname: Furriel
mesh_terms: D002241:Carbohydrates / Q000737:chemistry*; D004795:Enzyme Stability; D005285:Fermentation; D005374:Filtration; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008970:Molecular Weight; D010209:Paper; D020037:Sordariales / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature; D018505:Waste Management; D001617:beta-Glucosidase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D002241:Carbohydrates; D008670:Metals; D001617:beta-Glucosidase