| Summary: | This thesis is focused in the identification and improvement of lipases for biotechnological application. The importance of lipases is increasing in several industries. However, the commercial use of lipases is still a drawback in the economics of the lipase-based industrial applications. There are many tools for improving and adapting the enzyme properties to the desired requirements of a process that could lead lipase catalysis through a cost-effective process. In this context, the main objective of this work was: “To characterize, express and to improve novel bacterial lipases for sustainable industrial processes”. The first activity done was to explore and to characterize a new esterase from P. barcinonensis. It was isolate from P. barcinonensis the gene corresponding to Est23, and its cloning in a proper vector to perform expression and purification for biochemical characterization. Est23 showed preference for mid-chain substrates and having maximum activity at 37 °C and pH 7. It also includes in silico analysis of the 3D model structure and phylogeny. Moreover, a phylogenetic tree was constructed to assign Est23 to one of the bacterial hydrolase families described by Arpingy and Jaeger. Est23 could not be assigned to any bacterial hydrolases described till that moment, suggesting that Est23 could be part of a new group of bacterial lipases. Because Est23 displays a GGG(A)X-type putative oxyanion hole, widely described as a motif involved in tertiary alcohol resolution, the ability of Est23 for conversion and resolution of tertiary alcohols was evaluated. However, no conversion was detected using the esters linalyl and terpinyl acetate alcohols as substrates. To improve LipR activity by protein engineering LipR was then desired. LipR was isolated from Rhodococcus sp. strain CR-53 in a previous and showed an unusual fungal-like oxyanion-hole never found among bacterial lipases, close to the Y-type oxyanion hole described for Candida antartica lipase A (CalA), a lipase widely used in industry. In order to ...
|
|---|