SAXS Studies of the Endoglucanase Cel12A from Gloeophyllum trabeum Show Its Monomeric Structure and Reveal the Influence of Temperature on the Structural Stability of the Enzyme
Lis S. Miotto,
Caio V. dos Reis,
Mario de Oliveira Neto,
Igor Polikarpov
Affiliations
Lis S. Miotto
Grupo de Biotecnologia Molecular, Instituto de Física de São Carlos, Universidade de São Paulo, Av. Trabalhador São-Carlense 400, Arnold Schimidt, São Carlos, SP 13566-590, Brazil
Caio V. dos Reis
Grupo de Biotecnologia Molecular, Instituto de Física de São Carlos, Universidade de São Paulo, Av. Trabalhador São-Carlense 400, Arnold Schimidt, São Carlos, SP 13566-590, Brazil
Mario de Oliveira Neto
Departamento de Física e Biofísica, Instituto de Biociências de Botucatu, Universidade Estadual Paulista, Distrito de Rubião Júnior, S/N, Botucatu, SP 18618-970, Brazil
Igor Polikarpov
Grupo de Biotecnologia Molecular, Instituto de Física de São Carlos, Universidade de São Paulo, Av. Trabalhador São-Carlense 400, Arnold Schimidt, São Carlos, SP 13566-590, Brazil
Endoglucanases are key enzymes applied to the conversion of biomass aiming for second generation biofuel production. In the present study we obtained the small angle X-ray scattering (SAXS) structure of the G. trabeum endo-1,4-β-glucanase Cel12A and investigated the influence of an important parameter, temperature, on both secondary and tertiary structure of the enzyme and its activity. The CD analysis for GtCel12A revealed that changes in the CD spectra starts at 55 °C and the Tm calculated from the experimental CD sigmoid curve using the Boltzmann function was 60.2 ± 0.6 °C. SAXS data showed that GtCel12A forms monomers in solution and has an elongated form with a maximum diameter of 60 ± 5 Å and a gyration radius of 19.4 ± 0.1 Å as calculated from the distance distribution function. Kratky analysis revealed that 60 °C is the critical temperature above which we observed clear indications of denaturation. Our results showed the influence of temperature on the stability and activity of enzymes and revealed novel structural features of GtCel12A.
