title: Biochemical and biophysical properties of a metagenome-derived GH5 endoglucanase displaying an unconventional domain architecture.
abstract: Endoglucanases are key enzymes in the degradation of cellulose, the most abundant polymer on Earth. The aim of this work was to perform the biochemical and biophysical characterization of CelE2, a soil metagenome derived endoglucanase. CelE2 harbors a conserved domain from glycoside hydrolase family 5 (GH5) and a C-terminal domain with identity to Calx-beta domains. The recombinant CelE2 displayed preference for hydrolysis of oat beta-glucan, followed by lichenan and carboxymethyl cellulose. Optimum values of enzymatic activity were observed at 45°C and pH 5.3, and CelE2 exhibited considerable thermal stability at 40°C for up to 360min. Regarding the cleavage pattern on polysaccharides, the release of oligosaccharides with a wide degree of polymerization indicated a characteristic of endoglucanase activity. Furthermore, the analysis of products generated from the cleavage of cellooligosaccharides suggested that CelE2 exhibited transglycosylation activity. Interestingly, the presence of CaCl2positively affect CelE2, including in the presence of surfactants. SAXS experiments provided key information on the effect of CaCl2on the stability of CelE2 and dummy atom and rigid-body models were generated. To the best of our knowledge this is the first biochemical and biophysical characterization of an endoglucanase from family GH5 displaying this unconventional modular organization.
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil; Departamento de Bioquímica, Instituto de Biologia (IB), Universidade Estadual de Campinas (UNICAMP), R. Monteiro Lobato, 255 - Cidade Universitária, Campinas, São Paulo, Brazil. Electronic address: agnescristinap@gmail.com.
authors.forename: Agnes C
authors.initials: AC
authors.lastname: Pimentel
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil. Electronic address: gabriela.ematsu@bioetanol.org.br.
authors.forename: Gabriela C G
authors.initials: GCG
authors.lastname: Ematsu
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil. Electronic address: marcelovliberato@gmail.com.
authors.forename: Marcelo V
authors.initials: MV
authors.lastname: Liberato
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil; Departamento de Bioquímica, Instituto de Biologia (IB), Universidade Estadual de Campinas (UNICAMP), R. Monteiro Lobato, 255 - Cidade Universitária, Campinas, São Paulo, Brazil. Electronic address: douglas.paixao@bioetanol.org.br.
authors.forename: Douglas A A
authors.initials: DAA
authors.lastname: Paixão
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil; Departamento de Bioquímica, Instituto de Biologia (IB), Universidade Estadual de Campinas (UNICAMP), R. Monteiro Lobato, 255 - Cidade Universitária, Campinas, São Paulo, Brazil. Electronic address: jpcairo@gmail.com.
authors.forename: João Paulo L
authors.initials: JPL
authors.lastname: Franco Cairo
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil. Electronic address: fernanda.mandelli@bioetanol.org.br.
authors.forename: Fernanda
authors.initials: F
authors.lastname: Mandelli
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil; Programa de Pós Graduação em Biociências e Tecnologia de Produtos Bioativos (BTPB), Instituto de Biologia (IB) - CP 6109, Universidade Estadual de Campinas (UNICAMP), 13083-970, Campinas, SP, Brazil. Electronic address: robson.tramontina@gmail.com.
authors.forename: Robson
authors.initials: R
authors.lastname: Tramontina
authors.affiliation: Departamento de Física e Biofísica, Instituto de Biociências de Botucatu, UNESP Univ Estadual Paulista, Distrito de Rubião Jr. s/n, Botucatu, SP, Brazil. Electronic address: cesargandin@gmail.com.
authors.forename: César A
authors.initials: CA
authors.lastname: Gandin
authors.affiliation: Departamento de Física e Biofísica, Instituto de Biociências de Botucatu, UNESP Univ Estadual Paulista, Distrito de Rubião Jr. s/n, Botucatu, SP, Brazil. Electronic address: mario.neto@ibb.unesp.br.
authors.forename: Mario
authors.initials: M
authors.lastname: de Oliveira Neto
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil. Electronic address: fmsquina@gmail.com.
authors.forename: Fabio M
authors.initials: FM
authors.lastname: Squina
authors.affiliation: Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, CEP 13083-970, Campinas, São Paulo, Brazil. Electronic address: thabata.alvarez@up.edu.br.
authors.forename: Thabata M
authors.initials: TM
authors.lastname: Alvarez
mesh_terms: D055592:Biophysical Phenomena; D002480:Cellulase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004492:Edetic Acid / Q000494:pharmacology; D006031:Glycosylation; D054892:Metagenome; D008670:Metals / Q000494:pharmacology; D010802:Phylogeny; D011489:Protein Denaturation; D000072417:Protein Domains; D013379:Substrate Specificity; D013501:Surface-Active Agents / Q000494:pharmacology; D013696:Temperature
chemical_list: D008670:Metals; D013501:Surface-Active Agents; D004492:Edetic Acid; D002480:Cellulase
keywords: Calx-beta domain; Glycoside hydrolase family 5; Lignocellulosic biomass