Functional and structural studies of enzyme class endoglucanases GHF45 aiming for ...
| Grant number: | 15/18149-6 |
| Support type: | Scholarships in Brazil - Post-Doctorate |
| Effective date (Start): | November 01, 2015 |
| Effective date (End): | December 16, 2015 |
| Field of knowledge: | Biological Sciences - Biophysics |
| Principal Investigator: | Richard John Ward |
| Grantee: | Raquel Fonseca Maldonado |
| Home Institution: | Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (FFCLRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil |
| Associated research grant: | 10/18850-2 - Identification, characterization and engineering of plant cell wall degrading enzymes, AP.TEM |
Abstract Enzymatic hydrolysis of cellulose has attracted significant interest in the last years, and can be seen as a key enabling technology within the goal for the viable production of ethanol from lignocellulosic biomass. Despite the recent advances in cellulose saccharification, the availability of effective enzymes is still a limiting factor in reducing the cost of second generation bioethanol. Many enzymes involved in cellulose hydrolysis have a multidomain structure, containing a catalytic domain and one or more carbohydrate binding modules (CBM). Several studies have reported that CBMs facilitate substrate binding and access to the active site of the catalytic domain, promoting substrate specificity and stability of the enzyme/substrate complex. However, the conformational changes between the catalytic domain and the CBM in the presence of natural substrates remains unknown, and such knowledge is necessary to completely understand the complex mechanism of biomass hydrolysis by these multi-domain enzymes. In this project we will use Electron Magnetic Resonance (EMR) and the Site Directed Spin Labeling (SDSL) to study the dynamics of the catalytic domain of a GH5 endoglucanase from Bacillus subtilis fused with the CBM11 from Clostridium thermocellum in the presence of a natural substrate. With this goal, spin site directed mutants will be constructed and variations in their intramolecular distances will be measured and structural models will be constructed. To the best of our knowledge, this is the first time EMR has been used to study interdomain dynamics in glycosyl hydrolases, and the results will contribute to our understanding of the catalytic mechanism of these enzymes. | |