Degradação de penas por Bacillus sp. CL18: propriedades da protease bruta produzida durante cultivos submersos

Abstract

Keratin-rich wastes are generated in large quantities by agro-industrial activities, such as feathers from the poultry slaughterhouses, that need to be adequately disposed. In this sense, biotechnology emerges as a management and value-adding strategy for these wastes. This study aimed to evaluate the feather-degrading potential of the bacterial isolate Bacillus sp. CL18, to characterize the crude protease produced during cultivations on Feather Broth, and to verify its potential for the hydrolysis of various protein substrates. This strain degraded 95% of the feathers in the medium after 4 days of cultivation, resulting in increased soluble protein concentrations. The bacterium produced extracellular proteases for this process, reaching maximal values at the 4 th day of cultivation. This crude protease showed temperature and pH optima at 55 °C and 8.0, respectively. Enzyme thermal stability was not affected at 45 °C, while pre-incubations at 50-60 °C resulted in diminished residual activities, with half-life values decreasing as the temperature increased. The enzyme kinetics followed Michaelis- Menten equation. Cobalt, copper, iron and zinc salts, and the reducing agent β- mercaptoethanol resulted in decreased enzymatic activity, whereas calcium and magnesium salts, and the non-ionic detergents Triton X-100 and Tween 20, favored protease activity. Manganese salt and ethanol in the reaction media had no apparent effects on enzyme activity. Protease inhibitors PMSF and EDTA strongly decreased the crude protease activity, indicating the presence of serine and metalloproteases. The crude protease preferentially hydrolyzed casein and soy protein isolate. From these results, Bacillus sp. CL18 could be employed in feather degradation, and the crude protease produced during the cultures might be potentially applied in protein hydrolysis, aiming to obtain useful products.