Investigation of lignocellulosic degradation by the termite Coptotermes gestroi complementary to glycoside hydrolases aiming at the production of bioproducts

Author(s)

João Paulo Lourenço Franco Cairo

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia
• Defense date: 2016-08-31

Examining board members

Fabio Marcio Squina; Ednildo de Alcântara Machado; Marcos Silveira Buckeridge; Celso Eduardo Benedetti; André Ricardo de Lima Damásio

• Advisor: Fabio Marcio Squina

Abstract

Termites are social insects living in colonies and dividing the labor among the castes. This insect has its diet based on the consumption of lignocelluloses. The worker caste is responsible for feeding the colony, which turn the termites into on of the smaller bioreactor on Earth, and one of the most efficient organisms in the decomposition of lignocelluloses. The degradation of this material achieve approximately 75-90% efficiency and the lignin is modified. The digestion occurs in the termite gut, which occurs the secretion of carbohydrateactive enzymes (CAZymes), such as glycoside hydrolases (cellulases and hemicellulases). These enzymes are secreted in the fore- and midgut, and also by the symbionts, such as protozoa and bacteria, in the hindgut. However, these cellulases have been described in the literature as inefficient into degrading recalcitrant lignocellulose. Thus, the aim of this thesis was to investigate the occurrence of complementary mechanisms to glycoside hydrolases in the digestome the termite Coptotermes gestroi, mainly redox mechanisms, which could explain the efficient lignocelluloses degradation by termites. Thus, two endogenous glycoside hydrolases from C. gestroi, an endo-1,4-â-glucosidase CgEG-1 (GHF-9) and a â-1,4- glucosidase CgBG-1 (GHF-1) were functionally and structurally characterized. The results showed that the two enzymes act in synergy in the C. gestroi¿s digestome, exhibiting a high degree of synergy in the degradation of polysaccharides and complex lignocellulose. However, the level of biomass saccharification was very low, in comparison to saccharification yields using fungal cellulases. Thus, aiming to explore the potential of CAZymes in C. gestroi, analysis of metatranscriptomic and metaproteomics were performed in the worker and soldier castes. The results revealed that both castes had a similar repertoire of CAZymes, including from the Auxiliaries Activities (AAs) group (laccases, and glucose oxidases) and oxidoreductases related to Pro-oxidation, Antioxidant, and Detoxification (PAD) processes. In addition, genes and enzymes classified as CAZy and PAD had higher abundance in the worker caste, suggesting a role of theses enzymes in lignocellulose breakdown. Subsequently, a transcriptome analysis of C. gestroi¿s workers, fed on different sugarcane bagasses, have revealed that the expression of genes encoding superoxide dismutases, a PAD enzyme, was modulated by the diet. Thus, the endogenous CGSOD-1 gene was cloned and the protein CgSOD-1 was functional and biologically characterized. CgSOD-1 is a typical SOD, showing synergy with CgEG-1 in the hydrolysis of â-glucan. CgSOD-1 was immunolocalized in the fore- and midgut of C. gestroi, as well as CgEG-1, thus, suggesting the role of this SOD in the digestome. Biochemical analyzes also revealed that CgSOD-1 acts on the cleavage of polysaccharides, throughout the generation of reactive oxygen species. Moreover, CgSOD-1 was used to supplement cellulolytic cocktails, revealing a 30% increase in the biomass saccharification. In conclusion, this thesis has shown that the glycoside hydrolases produced by the lower termite C. gestroi are not efficient in the degradation of complex lignocellulose, moreover, it was evidenced that C. gestroi synthesized auxiliary enzymes to glycoside hydrolases, primarily oxidoreductases, which could act in the conversion of lignocellulosic biomass in the digestome of this insect (AU)

• FAPESP's process: 11/20977-3 - Investigation of plant biomass conversion in termite Coptotermes gestroi complementary to glycosyl hydrolases aiming at bioproducts formation derived from lignocellulosic biomass.
• Grantee: João Paulo Lourenço Franco Cairo
• Support Opportunities: Scholarships in Brazil - Doctorate