Caracterización de una enzima celulasa obtenida mediante reconstrucción ancestral de secuencias

Abstract

Modern enzymes are biological alternatives to the degradation of cellulosic biomass derived from agro-industrial waste, facilitating the production of high value-added products such as biofuels. However, the properties of current enzymes are not always suitable for their application in industrial environments. Ancestral sequence reconstruction is an in-silico technique that allows the deduction of ancient protein sequences from existing ones, based on the premise that on the primitive earth enzymes were more stable. We proceeded to the reconstruction of an ancestral enzyme from representative cellulases of family 6 of glycosyl hydrolases, with the expectation of obtaining improved properties and proceeding to their characterization. For the prediction of optimal conditions, the statistical method known as response surface methodology was used, considering pH and reaction temperature as variables. The results revealed that ancestral cellulase acts on the crystalline structure of cellulose. In addition, the determination of optimum conditions showed that cellulase exhibits higher activity at pH 8.6, compared to its current counterparts, which exhibit higher catalysis under acidic conditions. Also, the predicted temperature exceeds 76 degrees Celsius, which when compared to its counterparts that present temperatures up to a maximum of 60 degrees Celsius, suggesting a higher thermostability.