Caracterización de dos glicosiltransferasas productoras de oligosacáridos prebióticos de las levaduras phaffia rhodozyma y rhodotorula dairenensis

Resumo

Summary humans are one the most complex ecosystems formed by a large microbiome that is composed by ~1014 microoganisms versus only 1013 of human cells. The concept of human microbiome was defined by joshua lederberg as the communities of microorganisms that exist on the surface and inside the human body. They include mainly bacteria, but also fungi and archaea. Prebiotic oligosaccharides have been recognized as functional food ingredients since they selectively stimulate the growth or activity of potentially health-promoting colonic bacteria (lactobacilli and bifidobacteria). The byproducts generated by these microorganisms during the metabolism of prebiotics exert a beneficial effect on human health, contributing to the improvement of the immune response and potentially reducing the risk of acquiring several diseases (cancer, osteoporosis, cardiovascular disease or obesity, among others). In this project, two glycosylhydrolases from rhodotorula dairenensis y phaffia rhodozyma yeasts, which are capable of synthesizing oligosaccharides with potential prebiotics properties, have been studied. The ß-fructofuranosidase (ec 3.2.1.26) from rhodotorula dairenensis catalyses the release of fructose from the nonreducing termini of various ß-d-fructofuranoside substrates. As well as hydrolyzing sucrose, this enzyme catalyses the synthesis of short-chain fructooligosaccharides (fos), in which one or more fructosyl moieties are linked to the sucrose skeleton. The enzyme synthesizes basically a mixture of the prebiotics 6-kestose, neokestose and 1-kestose. The enzyme was purified, and it is composed of four subunits in which the monomer has a molecular weight of 172 kda. The ¿-glucosidase (ec 3.2.1.20) from phaffia rhodozyma catalyzes the release of glucose from the nonreducing termini of various substrates including ¿-(1¿4) bonds. This enzyme is able to produce a mixture of isomaltoligosaccharides with ¿-(1¿4) and ¿-(1¿6) bonds, which modulates the growth of various intestinal bacteria. The ¿-glucosidase gene was characterized and heterologous expressed in saccharomyces cerevisiae. The analysed sequence has 5.643 pb, with 968 pb as promotor, 4.175 pb as codifing sequece and 500 pb downstream of the stop codon. The gene also has 13 introns and has the same organization between different strains. The enzyme was classified within family 31 of the glycosylhydrolases. Palabras claves: Alfaglucosidasa, invertasa, rhodotorula, phaffia rhodozyma, prebioticos, oligosacaridos, imos, fos