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BACKGROUND
Various neutral and alkaline peptidases are commercially available for use in protein hydrolysis under neutral to alkaline conditions. However, the hydrolysis of proteins under acidic conditions by applying fungal aspartic peptidases (FAPs) has not been investigated in depth so far. The aim of this study, thus, was to purify a FAP from the commercial enzyme preparation, ROHALASE® BXL, determine its biochemical characteristics, and investigate its application for the hydrolysis of food and animal feed proteins under acidic conditions.
RESULTS
A Trichoderma reesei derived FAP, with an apparent molecular mass of 45.8 kDa (sodium dodecyl sulfate–polyacrylamide gel electrophoresis; SDS-PAGE) was purified 13.8-fold with a yield of 37% from ROHALASE® BXL. The FAP was identified as an aspartate protease (UniProt ID: G0R8T0) by inhibition and nano-LC-ESI-MS/MS studies. The FAP showed the highest activity at 50°C and pH 4.0. Monovalent cations, organic solvents, and reducing agents were tolerated well by the FAP. The FAP underwent an apparent competitive product inhibition by soy protein hydrolysate and whey protein hydrolysate with apparent Ki-values of 1.75 and 30.2 mg*mL−1, respectively. The FAP showed promising results in food (soy protein isolate and whey protein isolate) and animal feed protein hydrolyses. For the latter, an increase in the soluble protein content of 109% was noted after 30 min.
CONCLUSION
Our results demonstrate the applicability of fungal aspartic endopeptidases in the food and animal feed industry. Efficient protein hydrolysis of industrially relevant substrates such as acidic whey or animal feed proteins could be conducted by applying fungal aspartic peptidases. © 2022 Society of Chemical Industry.
The cellulase-producing Trichoderma reesei strain RL-P37 exhibits significant potential, yielding 7.3 g/L of cellulase in 241 hours. Microscopic investigations reveal a link between spore formation and enzyme production, suggesting the need for research into the intricate relationship between enzyme production, stress responses, and the nutritional prerequisites of fungi. Comparatively, the use of sodium hydroxide (NaOH) treatment, as opposed to water treatment, results in the reduction of micronutrient content and carbon source extraction as filtrate. Despite these challenges, research by He et al. (2021) highlights NaOH's efficiency in cellulose extraction from plant-based sources. Using NaOH pretreatment can be proven as effective by designing a proper cultivation method. The selection of inducers for enzyme induction gains importance, with soluble inducers, as emphasized by Zhang et al. (2022), exhibiting superior effectiveness. Hence, adopting soluble inducers in designing cultivation methods for improved enzyme production in shaking flasks is recommended. Enzymatic treatment of bio-waste, as outlined by Hu et al. (2021), shows promise in augmenting essential component content by breaking down plant cell walls and intercellular compartments. However, the feasibility of using an artificial bio-waste medium for cultivating Trichoderma reesei is questioned. Investigating the impact of micronutrient levels, particularly the inhibitory role of zinc, on fungal growth becomes essential. These findings underscore the necessity for ongoing research and optimization in cellulase production, emphasizing both strain productivity and cultivation methodologies.