Plant cell wall architecture guided design of CBM3-GH11 chimeras with enhanced xylanase activity using a tandem repeat left-handed beta-3-prism scaffold

Autor(es)

Pinheiro, P. Matheus ^[1] ; Reis, A. G. Renata ^[2] ; Dupree, Paul ^[3] ; Ward, J. Richard ^[1]

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Dept Quim, Fac Filosofia Ciencias & Letras Ribeirao Preto, Av Bandeirantes 3900, Bloco 18, Sala 8, BR-14040901 Ribeirao Preto, SP - Brazil

^[2] Univ Sao Paulo, Dept Quim & Fis, Fac Ciencias Farmaceut, BR-14040901 Ribeirao Preto, SP - Brazil

^[3] Univ Cambridge, Dept Biochem, Cambridge CB2 1QW - England

• Tipo de documento: Artigo Científico
• Fonte: COMPUTATIONAL AND STRUCTURAL BIOTECHNOLOGY JOURNAL; v. 19, p. 1108-1118, 2021.
• Citações Web of Science: 0

Resumo

Effective use of plant biomass as an abundant and renewable feedstock for biofuel production and biorefinery requires efficient enzymatic mobilization of cell wall polymers. Knowledge of plant cell wall composition and architecture has been exploited to develop novel multifunctional enzymes with improved activity against lignocellulose, where a left-handed beta-3-prism synthetic scaffold (BeSS) was designed for insertion of multiple protein domains at the prism vertices. This allowed construction of a series of chimeras fusing variable numbers of a GH11 beta-endo-1,4-xylanase and the CipA-CBM3 with defined distances and constrained relative orientations between catalytic domains. The cellulose binding and endoxylanase activities of all chimeras were maintained. Activity against lignocellulose substrates revealed a rapid 1.6- to 3-fold increase in total reducing saccharide release and increased levels of all major oligosaccharides as measured by polysaccharide analysis using carbohydrate gel electrophoresis (PACE). A construct with CBM3 and GH11 domains inserted in the same prism vertex showed highest activity, demonstrating interdomain geometry rather than number of catalytic sites is important for optimized chimera design. These results confirm that the BeSS concept is robust and can be successfully applied to the construction of multifunctional chimeras, which expands the possibilities for knowledge-based protein design. (C) 2021 The Authors. Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology. (AU)

• Processo FAPESP: 12/01066-2 - Desenho de enzimas multifuncionais que combinam a hidrólise de B-Glucanos e xilanos para avaliação da conformação destes polissacarídeos na parede celular de plantas
• Beneficiário: Matheus Pinto Pinheiro
• Modalidade de apoio: Bolsas no Brasil - Pós-Doutorado
• Processo FAPESP: 16/24139-6 - Desenho e Caracterização de Enzimas Xilanolíticas Multifuncionais
• Beneficiário: Richard John Ward
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 14/13956-8 - Investigação da arquitetura de polissacarídeos na parede celular de plantas de diferentes matérias-primas lignocelulósicas
• Beneficiário: Matheus Pinto Pinheiro
• Modalidade de apoio: Bolsas no Exterior - Estágio de Pesquisa - Pós-Doutorado
• Processo FAPESP: 14/50884-5 - INCT 2014: Instituto Nacional de Ciência e Tecnologia do Bioetanol
• Beneficiário: Marcos Silveira Buckeridge
• Modalidade de apoio: Auxílio à Pesquisa - Temático