title: Secretory expression and characterization of two hemicellulases, xylanase, and β-xylosidase, isolated from Bacillus subtilis M015.
abstract: Microbial hydrolysis of lignocellulosic biomass is becoming increasingly important for the production of renewable biofuels to address global energy concerns. Hemicellulose is the second most abundant lignocellulosic biopolymer consisting of mostly xylan and other polysaccharides. A variety of enzymes is involved in complete hydrolysis of xylan into its constituent sugars for subsequent biofuel fermentation. Two enzymes, endo-β-xylanase and β-xylosidase, are particularly important in hydrolyzing the xylan backbone into xylooligosaccharides and individual xylose units. In this study, we describe the cloning, expression, and characterization of xylanase and β-xylosidase isolated from Bacillus subtilis M015 in Escherichia coli. The genes were identified to encode a 213 amino acid protein for xylanase (glycoside hydrolase (GH) family 11) and a 533 amino acid protein for β-xylosidase (GH family 43). Recombinant enzymes were produced by periplasmic-leaky E. coli JE5505 and therefore secreted into the supernatant during growth. Temperature and pH optima were determined to be 50 °C and 5.5-6 for xylanase and 35 °C and 7.0-7.5 for β-xylosidase using beech wood xylan and p-nitrophenyl-β-D-xylopyranoside as the substrates, respectively. We have also investigated the synergy of two enzymes on xylan hydrolysis and observed 90 % increase in total sugar release (composed of xylose, xylobiose, xylotriose, and xylotetraose) for xylanase/β-xylosidase combination as opposed to xylanase alone.
authors.affiliation: Department of Chemical Engineering, University of Michigan, 3074 H.H.Dow Buil., 2300 Hayward St, Ann Arbor, MI, 48109, USA.
authors.forename: Alison L
authors.initials: AL
authors.lastname: Banka
authors.forename: Saadet Albayrak
authors.initials: SA
authors.lastname: Guralp
authors.forename: Erdogan
authors.initials: E
authors.lastname: Gulari
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics; D043364:Endo-1,4-beta Xylanases / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D006026:Glycoside Hydrolases / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D006868:Hydrolysis; D008031:Lignin / Q000737:chemistry*; D011994:Recombinant Proteins; D014995:Xylosidases / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; C036909:lignocellulose; D008031:Lignin; D006026:Glycoside Hydrolases; D014995:Xylosidases; C023305:hemicellulase; D043364:Endo-1,4-beta Xylanases