Expression and purification of recombinants xylanolitics enzymes and study of their activities against sugar cane extracted arabinoxylans

• Grant number: 18/25664-2
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• Start date: June 01, 2019
• End date: March 31, 2022
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Richard John Ward
• Grantee: Carolina Victal Garbelotti
• Host Institution: Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (FFCLRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated research grant: 14/50884-5 - INCT 2014: National Institute of Science and Technology of Bioethanol, AP.TEM

Abstract

The search for methods that use lignocellulosic biomass for production of second generation ethanol has lead to the study of enzymes able to hydrolyse this substrate and provide fermentable sugars. However, the heterogenic and recalcitrant composition of lignocellulosic biomass require a group of enzymes acting synergistically for its complete hydrolysis. This works aims to express, purify and characterize the activity of the enzymes Feruloyl Esterase (Fae), Acetyl Xylan Esterase (Axe), Arabinofuranosidase (Ara), Xylanase (Xyl) and alpha-Glucuronidase (AGlu), which have as substrate different glycosidic bonds on the polysaccharide arabinoxylan, part of that complex feedstock. pET-28 plasmids containing the enzymes insert will be transformed in Escherichia coli strains. The enzymes will be purified by affinity chromatography and used in the activity essays, alone and combined, against wheat and sugar cane arabinoxylan. Mass spectrometry will be used for detection of structures released after enzymatic treatment and imaging of the polymer surface, thus aiming to understand the architecture of the plant cell wall and the mechanism of synergistic action of lignocellulolytic enzymes and the possibility of elaborating a mix of enzymes that could work synergistically and efficiently hydrolyse lignocellulosic biomass. (AU)

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)

BARRETO, MATHEUS QUINTANA; GARBELOTTI, CAROLINA VICTAL; SOARES, JESSICA DE MOURA; GRANDIS, ADRIANA; BUCKERIDGE, MARCOS SILVEIRA; LEONE, FRANCISCO ASSIS; WARD, RICHARD JOHN. Xylose isomerase from Piromyces sp. E2 is a promiscuous enzyme with epimerase activity. Enzyme and Microbial Technology, v. 166, p. 9-pg., 2023-03-24. (16/24139-6, 14/50884-5, 18/25664-2)

GARBELOTTI, CAROLINA V.; BULMER, GREGORY S.; WARD, RICHARD J.; VAN MUNSTER, JOLANDA M.. The feruloyl esterase from Thermobacillus xylanilyticus shows broad specificity for processing pre-biotic feruloylated xylooligosaccharides at high temperatures. Food Chemistry, v. 405, p. 8-pg., 2023-03-30. (14/50884-5, 18/14030-2, 18/25664-2)

TRINCA, VITOR; CARLI, SIBELI; ULIANA, JOAO VITOR CARDOSO; GARBELOTTI, CAROLINA VICTAL; DA SILVA, MARIANA MENDES; KUNES, VITOR; MELEIRO, LUANA PARRAS; BRANCINI, GUILHERME THOMAZ PEREIRA; MENZEL, FRANK; ANDRIOLI, LUIZ PAULO MOURA; et al. Biocatalytic potential of Pseudolycoriella CAZymes (Sciaroidea, Diptera) in degrading plant and cell wall. ISCIENCE, v. 26, n. 4, p. 27-pg., 2023-04-21. (14/14318-5, 20/05636-4, 16/17582-0, 16/24139-6, 16/25325-8, 17/13734-3, 18/25664-2)

Academic Publications

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

GARBELOTTI, Carolina Victal. Expression and purification of recombinants xylanolytic enzymes and study of their activities against sugar cane extracted arabinoxylans. 2022. Doctoral Thesis - Universidade de São Paulo (USP). Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (PCARP/BC) Ribeirão Preto.