Bioprospecting and construction of hemicellulases for the production of xylose from hydrolyzate licor pentoses and their use in fermentative xylitol production

• Grant number: 13/10443-7
• Support Opportunities: Scholarships in Brazil - Master
• Start date: August 01, 2013
• End date: July 31, 2015
• Field of knowledge: Biological Sciences - Biochemistry - Enzymology
• Principal Investigator: Hélia Harumi Sato
• Grantee: José Alberto Diogo
• Host Institution: Faculdade de Engenharia de Alimentos (FEA). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil

Abstract

The plant biomasses are composed mainly of cellulose, hemicellulose and lignin. Cellulose and hemicellulose can be hydrolyzed to monomeric sugars (primarily xylose/C5 and glucose/C6) using cellulases and hemicellulases. These molecules are sources of raw material for important industrial products as xylitol and ethanol. However, the tasks of a process for enzymatic hydrolysis of biomass saccharification to sugars into a pipeline using low cost and high efficiency, today is a challenge due to the complexity of these polysaccharides along with its recalcitrant effects. Hemicellulases are enzymes of glycoside hydrolyses group which catalyze the hemicellulose hydrolysis, these enzymes acting synergistically together another cellulases and ligninases promoting the total and efficient hydrolysis of plant biomass. The hemicellulose is composed around of 30% of the polysaccharide in the biomass matrix of sugarcane bagass, however few studies are directed and focused to improve of saccharification (especially using hemicelulasic hydrolysis) of these specific hemi-polysaccharides to sugars and then its complete conversion to another value-added byproduct (bioethanol, biodiesel, organic acids and sugar as xylose and xylitol).In this sense, this project will be focused in the first step in the bioprospection of hemicellulolitics enzymes of microorganisms capable of growing in liquor pentoses from the hydrothermal pre-treatment of the crushed cane sugar. For this, from a library of thermophilic microorganisms with hemicellulolitic potential (previously detected as strains of Bacillus sp) will be screened exploring its ability to grow on solid medium containing liquor pentose (C5 fractions rich in sugars and xylo-oligomers). Once you select the best strain, will be performed the cloning, expression and characterization of an endo-xylanase (GH11) and a ²-xylosidase (GH43), key enzymes for xylose release from oligo-and polysaccharides. Isolated and characterized these best and efficient hemicellulases will be the construction of the quimeric enzyme (bifunctional enzyme), through molecular engineering this construction will be optimized for the hydrolysis of xylo-oligomers of pentoses present in this liquor. The xyloligomers of liquors will be hydrolyzed enzymatically enriched after this, the monosaccharides (mainly xylose) will be used to produce xylitol via fermentation using Scheffersomyces stipitis yeast (formely Pichia stipitis). The xylitol is one sugar alcohol with low caloric content and interesting properties nutraceutical. In parallel, this described system will be evaluated for its potential for the production of xylose and conversion of xylitol from free and immobilized yeasts (Scheffersomyces stipitis)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

DIOGO, JOSE A.; HOFFMAM, ZAIRA B.; ZANPHORLIN, LETICIA M.; COTA, JUNIO; MACHADO, CARLA B.; WOLF, LUCIA D.; SQUINA, FABIO; DAMASIO, ANDRE R. L.; MURAKAMI, MARIO T.; RULLER, ROBERTO. Development of a chimeric hemicellulase to enhance the xylose production and thermotolerance. Enzyme and Microbial Technology, v. 69, p. 31-37, FEB 2015. (11/14200-6, 13/10443-7, 08/58037-9)

DIOGO, JOSE ALBERTO; ZANPHORLIN, LETICIA MARIA; SATO, HELIA HARUMI; MURAKAMI, MARIO TYAGO; RULLER, ROBERTO. Molecular cloning, overexpression, purification and crystallographic analysis of a GH43-xylosidase from Bacillus licheniformis. ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS, v. 71, p. 4-pg., 2015-08-01. (13/10443-7, 13/13309-0, 14/07135-1)

DIOGO, JOSE ALBERTO; ZANPHORLIN, LETICIA MARIA; SATO, HELIA HARUMI; MURAKAMI, MARIO TYAGO; RULLER, ROBERTO. Molecular cloning, overexpression, purification and crystallographic analysis of a GH43-xylosidase from Bacillus licheniformis. ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS, v. 71, n. 8, p. 962-965, AUG 2015. (13/13309-0, 13/10443-7, 14/07135-1)

Academic Publications

(References retrieved automatically from State of São Paulo Research Institutions)

DIOGO, José Alberto. Bioprospection and construction of hemicellulases for the production of xylose from the xylooligosaccharides liquor and its use in the fermentative xylitol production. 2016. Master's Dissertation - Universidade Estadual de Campinas (UNICAMP). Faculdade de Engenharia de Alimentos Campinas, SP.