EXPRESSION AND CHARACTERIZATION OF AN ASPARTIC PROTEASE FROM SOLANUM TUBEROSUM IN KLUYVEROMYCES LACTIS

PEPE, ALFONSO; MARÍA G. GUEVARA.; TONON, CLAUDIA VIRGINIA; TITO, FLORENCIA ROCÍO; GUSTAVO DALEO

Salta

Congreso; LV Reunión Anual de la Sociedad Argentina de Bioquímica y Biología Molecular; 2019

Sociedad Argentina de Bioquímica y Biología Molecular

In last years, the gradual decrease of cow population, the increasing global demands for cheese, the ethical problems associated with the isolation of the animal rennet and the incidence of bovine spongiform encephalopathy (BSE) have reduced both, supply and demand for bovine rennet. Therefore, several researches have been focused on discover new natural milk-clotting enzymes from microorganism and plants that would successfully replace the calf rennet in cheese manufacture. Previously, we have isolated and characterized an aspartic protease from S. tuberosum named as StAP3. StAP3 is able to clot bovine milk and to degrade bovine casein subunits. In this work, we report the development of a new StAP3-derived produced in the generally regarded as safe (GRAS) yeast K. lactis. Using a stepwise optimization strategy?consisting of culture media screening, complemented with a chromatography in a pepstatin A- agarose column step? we successfully improve StAP3 production in K. lactis (rStAP3). As StAP3, rStAP3 has caseinolytic and milk clotting activity with an optimum MCA value in accordance with those used in the industrial manufacture of cheeses (pH 6 - 6.2 and temperature 40 - 42 ºC). These results, in terms of milk-clotting activity, suggest the suitability of rStAP3 to producing milk clots and the possibility of using these proteases in the artisanal and industrial cheese production.