Structure, computational and biochemical analysis of PcCel45A endoglucanase from Phanerochaete chrysosporium and catalytic mechanisms of GH45 subfamily C members

Autor(es):Mostrar menos -

Godoy, Andre S. ^[1] ; Pereira, Caroline S. ^[2] ; Ramia, Marina Paglione ^[1] ; Silveira, Rodrigo L. ^[2] ; Camilo, Cesar M. ^[3] ; Kadowaki, Marco A. ^[1] ; Lange, Lene ^[4] ; Busk, Peter K. ^[4] ; Nascimento, Alessandro S. ^[1] ; Skaf, Munir S. ^[2] ; Polikarpov, Igor ^[1]

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Sao Carlos Inst Phys, BR-13566590 Sao Paulo - Brazil

^[2] Univ Estadual Campinas, Inst Chem, BR-13084862 Sao Paulo - Brazil

^[3] Ctr Tecnol Canavieira, POB 162, BR-13400970 Sao Paulo - Brazil

^[4] Tech Univ Denmark, Dept Chem & Biochem Engn, Bldg 229, DK-2800 Lyngby - Denmark

• Tipo de documento: Artigo Científico
• Fonte: SCIENTIFIC REPORTS; v. 8, FEB 27 2018.
• Citações Web of Science: 4

Resumo

The glycoside hydrolase family 45 (GH45) of carbohydrate modifying enzymes is mostly comprised of beta-1,4-endoglucanases. Significant diversity between the GH45 members has prompted the division of this family into three subfamilies: A, B and C, which may differ in terms of the mechanism, general architecture, substrate binding and cleavage. Here, we use a combination of X-ray crystallography, bioinformatics, enzymatic assays, molecular dynamics simulations and site-directed mutagenesis experiments to characterize the structure, substrate binding and enzymatic specificity of the GH45 subfamily C endoglucanase from Phanerochaete chrysosporium (PcCel45A). We investigated the role played by different residues in the binding of the enzyme to cellulose oligomers of different lengths and examined the structural characteristics and dynamics of PcCel45A that make subfamily C so dissimilar to other members of the GH45 family. Due to the structural similarity shared between PcCel45A and domain I of expansins, comparative analysis of their substrate binding was also carried out. Our bioinformatics sequence analyses revealed that the hydrolysis mechanisms in GH45 subfamily C is not restricted to use of the imidic asparagine as a general base in the ``Newton's cradle{''} catalytic mechanism recently proposed for this subfamily. (AU)

• Processo FAPESP: 15/13684-0 - Estudos estruturais e funcionais de enzimas que participam na síntese e degradação de carboidratos complexos
• Beneficiário: Igor Polikarpov
• Modalidade de apoio: Auxílio à Pesquisa - Temático
• Processo FAPESP: 11/21608-1 - Identificação e caracterização de novas enzimas com potencial na digestão de biomassa lignocelulósica
• Beneficiário: Bruno Luan Soares Paula de Mello
• Modalidade de apoio: Bolsas no Brasil - Doutorado Direto
• Processo FAPESP: 10/52362-5 - Targeted analysis of microbial lignocellulolytic secretomes: a new approach to enzyme discovery
• Beneficiário: Igor Polikarpov
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 11/20505-4 - Duas classes importantes de glicosil hidrolases: estudos funcionais e análise estrutural
• Beneficiário: Marco Antonio Seiki Kadowaki
• Modalidade de apoio: Bolsas no Brasil - Pós-Doutorado