Engineering of an endopolygalacturonase by insertion of different Carbohydrate Binding Modules (CBMs): chimerogenesis and proximity effects

• Grant number: 17/13734-3
• Support Opportunities: Scholarships in Brazil - Doctorate
• Start date: February 01, 2018
• End date: October 31, 2020
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Richard John Ward
• Grantee: Sibeli de Carli
• Host Institution: Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (FFCLRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated scholarship(s): 19/26891-5 - Effects of the pectinolytic activity of endopolygacturonase-CBMs chimeras on the saccharification of different biomass feedstocks, BE.EP.DR

Abstract

The large surplus of biomass resulting from processing sugarcane for 1st generation ethanol production, can be used for cogeneration of electricity or for the production of 2nd generation ethanol. Strategies to reduce the costs related to the enzymatic hydrolysis of these lignocellulosic residues include the identification and heterologous expression of enzymes with interesting catalytic characteristics, the use of protein engineering to improve their catalytic performance, and finally, the formulation of more efficient enzymatic cocktails. The controlled deconstruction of the plant cell wall lignocellulosic matrix by enzymes is challenging and constitutes a major barrier to achieve high-yield production of bioethanol at competitive prices. It is known that the presence of pectins, one of the main components of the middle lamella in plants, hinders the hydrolysis of the plant cell wall and prevents the efficient release of sugars. With the goal of developing technologies to optimize the hydrolysis of this polysaccharide, we intend to evaluate the effects of the proximity of different polysaccharides present in the plant cell wall on the activity of an endopolygalacturonase (endoPG I), engineered by the addition of a series of Carbohydrate Binding Modules (CBMs). Protein chimeras will be constructed from the fusion of endoPG I and different CBMs with specific affinities to cellulose, xylan, pectin, ²-glucan and xyloglucan. Following a biochemical characterization using a range of analytic techniques, these chimeras will be used to supplement enzymatic cocktails for the hydrolysis of lignocellulosic residues, such as straw and sugarcane bagasse, with the goal of increasing the yields of fermentable sugars. (AU)

Scientific publications

(9)

(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)

CARLI, SIBELI; BUFFONI DE CAMPOS CARNEIRO, LARA APARECIDA; WARD, RICHARD JOHN; MELEIRO, LUANA PARRAS. Immobilization of a beta-glucosidase and an endoglucanase in ferromagnetic nanoparticles: A study of synergistic effects. Protein Expression and Purification, v. 160, p. 28-35, AUG 2019. (17/13734-3, 16/17582-0)

CARLI, SIBELI; SALGADO, JOSE CARLOS SANTOS; MELEIRO, LUANA PARRAS; WARD, RICHARD JOHN. ovalent Immobilization of Chondrostereum purpureum Endopolygalacturonase on Ferromagnetic Nanoparticles: Catalytic Properties and Biotechnological Applicatio. Applied Biochemistry and Biotechnology, v. 194, n. 2, SEP 2021. (19/17958-9, 14/50884-5, 17/13734-3, 19/21989-7, 16/17582-0)

FURTADO, GILVAN PESSOA; CARLI, SIBELI; MELEIRO, LUANA PARRAS; SANTOS SALGADO, JOSE CARLOS; WARD, RICHARD JOHN. Enhanced hydrolytic efficiency of an engineered CBM11-glucanase enzyme chimera against barley beta-D-glucan extracts. Food Chemistry, v. 365, DEC 15 2021. (16/17582-0, 16/24139-6, 17/13734-3, 14/50884-5, 19/21989-7)

CARLI, SIBELI; MELEIRO, LUANA PARRAS; WARD, RICHARD JOHN. Biochemical and kinetic characterization of the recombinant GH28 Stereum purpureum endopolygalacturonase and its biotechnological application. International Journal of Biological Macromolecules, v. 137, p. 469-474, SEP 15 2019. (17/13734-3, 16/17582-0)

SANTOS SALGADO, JOSE CARLOS; MELEIRO, LUANA PARRAS; CARLI, SIBELI; WARD, RICHARD JOHN. Glucose tolerant and glucose stimulated beta-glucosidases - A review. Bioresource Technology, v. 267, p. 704-713, NOV 2018. (17/13734-3, 14/50884-5, 16/17582-0)

DE CARVALHO, DANIELLA ROMANO; CARLI, SIBELI; MELEIRO, LUANA PARRAS; ROSA, JOSE CESAR; CAVALCANTE DE OLIVEIRA, ARTHUR HENRIQUE; JORGE, JOAO ATILIO; MELO FURRIEL, ROSA PRAZERES. A halotolerant bifunctional beta-xylosidase/alpha-L-arabinofuranosidase from Colletotrichum graminicola: Purification and biochemical characterization. International Journal of Biological Macromolecules, v. 114, p. 741-750, JUL 15 2018. (17/13734-3, 14/14415-0, 16/17582-0)

BUFFONI DE CAMPOS CARNEIRO, LARA APARECIDA; FUZO, CARLOS ALESSANDRO; MELEIRO, LUANA PARRAS; CARLI, SIBELI; BARRETO, MATHEUS QUINTANA; LOURENZONI, MARCOS ROBERTO; BUCKERIDGE, MARCOS SILVEIRA; WARD, RICHARD JOHN. Selective xyloglucan oligosaccharide hydrolysis by a GH31 alpha-xylosidase from Escherichia coli. Carbohydrate Polymers, v. 284, p. 12-pg., 2022-05-15. (12/24147-8, 17/13734-3, 16/24139-6, 16/17582-0, 14/50884-5, 16/15708-7)

CARLI, SIBELI; PARRAS MELEIRO, LUANA; SALGADO, JOSE CARLOS SANTOS; WARD, RICHARD JOHN. Synthetic carbohydrate-binding module-endogalacturonase chimeras increase catalytic efficiency and saccharification of lignocellulose residues. BIOMASS CONVERSION AND BIOREFINERY, v. N/A, p. 12-pg., 2022-05-03. (16/24139-6, 16/17582-0, 17/13734-3, 19/21989-7, 14/50884-5)

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)

CARLI, Sibeli de. Engineering of an endopolygalacturonase by insertion of different Carbohydrate Binding Modules (CBMs): Chimerogenesis and proximity effects. 2020. Doctoral Thesis - Universidade de São Paulo (USP). Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (PCARP/BC) Ribeirão Preto.