Resumo

It is estimated that 75% of the Earth's biosphere is in perennially cold environments representing a huge unexplored resource for identifying new enzymes with potential industrial application. Enzymes with activity at low temperatures have great biotechnological value as they are capable of efficiently catalyzing reactions at moderate or low temperatures, thereby reducing costs associated with heating steps. The aim of this work was to identify and characterize enzymes with potential industrial application on samples from King’s George Island, located in the Fildes Peninsula of Antarctica. With this aim, classical microbiological and functional metagenomics approaches were performed. For the classical microbiological approach, twelve samples were taken at different sites, serially diluted, plated on solid media previously reported for the culture of slow-growth microorganisms (R2A) and grown at 4 °C. Forty eight isolates were selected from each sample on the basis on their phenotype. The total of the selected isolates (576 isolates) were screened for the expression of some enzymatic activities. To this end, isolates were grown on R2A agar containing specific indicators to assess cellulolytic, lipolytic, proteolytic, antimicrobial, laccase, peroxidase or iron scavenging activities. For the functional metagenomics approach we selected one of the samples and we performed genome DNA extractions by using two different methods in order to avoid methodological bias. The extraction method was optimized in order to recover high quality DNA with the appropriate size for the construction of metagenomics library. From the 576 isolates, 30 gave positive activity for cellulase, 11 for laccase, 13 for lipase, 13 for protease, 9 for the production of antimicrobial compounds and 11 positive for iron scavenging. All positive clones were tested at different temperatures and with alternative substrates to select most promising clones. We obtained high quality DNAs, but the fragment sizes were not optimal for the construction of metagenomics library. Further extractions are being held. In summary, by using a classical microbiological approach we were able to identify many isolates from a cold environment with different enzymatic activities with potential industrial applications. The functional metagenomics approach will further complement this work by the identification of additionaly enzymatic activities from cultivable as well as uncultivable bacteria.