Multi-user equipament approved in grant 2013/13309-0: Beckman Coulter capillary electrophoresis systems with LIF detection system

• Grant number: 14/07135-1
• Support Opportunities: Multi-user Equipment Program
• Start date: June 01, 2014
• End date: May 31, 2016
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Mário Tyago Murakami
• Grantee: Mário Tyago Murakami
• Host Institution: Centro Nacional de Pesquisa em Energia e Materiais (CNPEM). Ministério da Ciência, Tecnologia e Inovação (Brasil). Campinas , SP, Brazil
• Associated research grant: 13/13309-0 - Studies of the structural and functional behavior of enzymes evolutionarily specialized in the degradation of plant biomass with potential biotechnological applications, AP.R
• EMU web page: Página do Equipamento Multiusuário não informada
• Type of equipment: Caracterização e Análises de Amostras - Proteínas/Ácidos nuclêicos - Eletroforese
• Manufacturer: Fabricante não informado
• Model: Modelo não informado

Abstract

The structure-function relationship studies of glycoside hydrolases contribute to unveil new strategies to improve thermal stability, specificity, and modulation - knowledge that can be used for the development of optimized enzymes for biotechnological processes as well as to assist the rational design of enzymes. However, studies regarding the structural determinants for the different modes of action of these enzymes on the substrates, which would be a next step in the investigations, are limited due to the lack of information regarding the products formed by enzyme action. Thus, the characterization of the cleavage products combined with site-directed mutagenesis and crystallographic studies would allow us to elucidate the molecular basis of the mode of action of these enzymes and provides a better understanding beyond what the current tools available allow. The main experiment that is routinely performed in this machine is the analysis of reaction products of glycoside hydrolases and for that, the products will be labeled with APTS, a fluorescent molecule. Hence, for detection of APTS-labeled products is strictly necessary coupling a module of excitation and fluorescence detection. A number of novel enzymes have been discovered by genome sequencing as well as new approaches such as metagenomics, and the understanding of products formed by glycoside hydrolases and their biochemical and kinetic behavior would be crucial to assess the biotechnological potential of these enzymes. The state-of-the-art studies on functional and structural aspects of GHs have been based in 3D structure elucidation along with a detailed functional characterization including necessarily the analysis of cleavage products of wild-type and mutant enzymes. With this equipment, a next level of understanding of the mechanistic of these enzymes will be achieved. The Automated Laboratory for Protein Crystallization (RoboLab) is already a reality in LNBio and has been largely contributing to the determination of new 3D structures that combined with the new Capillary Electrophoresis System will provide a formidable set of tools to assess the mode of action of glycosidic enzymes. This acquisition will extend and integrate the LNBio research in this field as well as other universities and institutions which could be interested. (AU)

Scientific publications

(7)

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

CRESPIM, ELAINE; ZANPHORLIN, LETICIA M.; DE SOUZA, FLAVIO H. M.; DIOGO, JOSE A.; GAZOLLA, ALEX C.; MACHADO, CARLA B.; FIGUEIREDO, FERNANDA; SOUSA, AMANDA S.; NOBREGA, FELIPE; PELLIZARI, VIVIAN H.; et al. A novel cold-adapted and glucose-tolerant GH1 beta-glucosidase from Exiguobacterium antarcticum B7. International Journal of Biological Macromolecules, v. 82, p. 375-380, JAN 2016. (13/13309-0, 14/07135-1)

SANTOS, CAMILA RAMOS; HOFFMAM, ZAIRA BRUNA; DE MATOS MARTINS, VANESA PEIXOTO; ZANPHORLIN, LETICIA MARIA; DE PAULA ASSIS, LEANDRO HENRIQUE; HONORATO, RODRIGO VARGAS; LOPES DE OLIVEIRA, PAULO SERGIO; RULLER, ROBERTO; MURAKAMI, MARIO TYAGO. Molecular Mechanisms Associated with Xylan Degradation by Xanthomonas Plant Pathogens. Journal of Biological Chemistry, v. 289, n. 46, p. 32186-32200, NOV 14 2014. (13/13309-0, 10/51890-8, 14/07135-1)

DOS SANTOS, CAMILA RAMOS; DE GIUSEPPE, PRISCILA OLIVEIRA; MOREIRA DE SOUZA, FLAVIO HENRIQUE; ZANPHORLIN, LETICIA MARIA; DOMINGUES, MARIANE NORONHA; SIQUEIRA PIROLLA, RENAN AUGUSTO; HONORATO, RODRIGO VARGAS; COSTA TONOLI, CELISA CALDANA; BUENO DE MORAIS, MARIANA ABRAHAO; DE MATOS MARTINS, VANESA PEIXOTO; et al. The mechanism by which a distinguishing arabinofuranosidase can cope with internal di-substitutions in arabinoxylans. BIOTECHNOLOGY FOR BIOFUELS, v. 11, AUG 11 2018. (13/13309-0, 14/17264-3, 15/26982-0, 14/07135-1)

ZANPHORLIN, LETICIA MARIA; DE GIUSEPPE, PRISCILA OLIVEIRA; HONORATO, RODRIGO VARGAS; COSTA TONOLI, CELISA CALDANA; FATTORI, JULIANA; CRESPIM, ELAINE; LOPES DE OLIVEIRA, PAULO SERGIO; RULLER, ROBERTO; MURAKAMI, MARIO TYAGO. Oligomerization as a strategy for cold adaptation: Structure and dynamics of the GH1 beta-glucosidase from Exiguobacterium antarcticum B7. SCIENTIFIC REPORTS, v. 6, MAR 31 2016. (13/13309-0, 14/07135-1)

DIOGO, JOSE ALBERTO; ZANPHORLIN, LETICIA MARIA; SATO, HELIA HARUMI; MURAKAMI, MARIO TYAGO; RULLER, ROBERTO. Molecular cloning, overexpression, purification and crystallographic analysis of a GH43-xylosidase from Bacillus licheniformis. ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS, v. 71, p. 4-pg., 2015-08-01. (13/10443-7, 13/13309-0, 14/07135-1)

DIOGO, JOSE ALBERTO; ZANPHORLIN, LETICIA MARIA; SATO, HELIA HARUMI; MURAKAMI, MARIO TYAGO; RULLER, ROBERTO. Molecular cloning, overexpression, purification and crystallographic analysis of a GH43-xylosidase from Bacillus licheniformis. ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS, v. 71, n. 8, p. 962-965, AUG 2015. (13/13309-0, 13/10443-7, 14/07135-1)

FURTADO, GILVAN P.; SANTOS, CAMILA R.; CORDEIRO, ROSA L.; RIBEIRO, LUCAS F.; DE MORAES, LUIZ A. B.; DAMASIO, ANDR R. L.; POLIZELI, MARIA DE LOURDES T. M.; LOURENZONI, MARCOS R.; MURAKAMI, MARIO T.; WARD, RICHARD J.. Enhanced xyloglucan-specific endo-beta-1,4-glucanase efficiency in an engineered CBM44-XegA chimera. Applied Microbiology and Biotechnology, v. 99, n. 12, p. 5095-5107, JUN 2015. (10/18850-2, 12/20549-4, 13/18910-3, 13/13309-0, 14/07135-1, 10/07133-8)