The keratinolytic bacteria Bacillus cytotoxicus as a source of novel proteases and feather protein hydrolysates with antioxidant activities

Abstract

Background Argentina’s geothermal areas are niches of a rich microbial diversity. In 2020, species of Bacillus cytotoxicus were isolated for the first time from these types of pristine natural areas. Bacillus cytotoxicus strains demonstrated the capability to grow and degrade chicken feathers with the concomitant production of proteases with keratinolytic activity, enzymes that have multitude of industrial applications. The aim of this research was to study the production of the proteolytic enzymes and its characterization. Also, feather protein hydrolysates produced during fermentation were characterized. Results Among the thermotolerant strains isolated from the Domuyo geothermal area (Neuquén province, Argentina), Bacillus cytotoxicus LT-1 and Oll-15 were selected and put through submerged cultures using feather wastes as sole carbon, nitrogen, and energy source in order to obtain proteolytic enzymes and protein hydrolysates. Complete degradation of feathers was achieved after 48 h. Zymograms demonstrated the presence of several proteolytic enzymes with an estimated molecular weight between 50 and > 120 kDa. Optimum pH and temperatures of Bacillus cytotoxicus LT-1 crude extract were 7.0 and 40 °C, meanwhile for Oll-15 were 7.0 and 50 °C. Crude extracts were inhibited by EDTA and 1,10 phenanthroline indicating the presence of metalloproteases. Feather protein hydrolysates showed an interesting antioxidant potential measured through radical-scavenging and Fe3+-reducing activities. Conclusion This work represents an initial approach on the study of the biotechnological potential of proteases produced by Bacillus cytotoxicus. The results demonstrated the importance of continuous search for new biocatalysts with new characteristics and enzymes to be able to cope with the demands in the market.

Innovative Artificial-Intelligence- Based Approach for the Biodegradation of Feather Keratin by Bacillus paramycoides, and Cytotoxicity of the Resulting Amino Acids

“…Generally, keratin-degrading efficiency is well reported in several bacteria, especially Bacillus spp., that can thrive on the keratinous biomass as the sole source of nitrogen, and carbon, indicating the secretion of protein-and keratin-degrading enzymes (Nnolim et al, 2020;Bhari et al, 2021;Cavello et al, 2021;Li, 2021). Bacillus spp.…”

“…has been known as the predominant bacteria group that mediates the degradation of keratin compounds through keratinases and, moreover, has gained much attention for keratinase production because of scale-up feasibility, easy handling, and high commercial applicability (Nnolim et al, 2020). The species include B. subtilis, B. aerius, B. pumilus, B. cytotoxicus, B. cereus, and B. lichenifomis (Cavello et al, 2021;Li, 2021). The latter is the first commercial keratinase preparation source (Bhari et al, 2021).…”

Keratinases as Versatile Enzymatic Tools for Sustainable Development

“…For in vitro food-protein hydrolysis, proteases with broad substrate specificity, from plant (ficin, bromelain, papain), animal (pepsin, chymotrypsin, trypsin) and microbial (proteinase K, collagenase, subtilisin A) sources, are needed [ 136 ]. As the representatives of proteolytic enzymes, keratinases can be potentially used for bioactive peptides production via enzymatic hydrolysis of keratins and other proteins, or by full cell fermentation of microbial media supplemented with poultry feather and non-keratin plant proteins [ 68 , 142 , 145 ].…”

“…Feather hydrolysates were also obtained by full-cell fermentation of Bacillus cytotoxicus LT-1 and B. cytotoxicus Oll-15 [ 68 ]. Minimal mineral media (2.48 g/L K 2 HPO 4 , 0.49 g/L NaH 2 PO 4 , 0.01 g/L MgCl 2 , 0.02 g/L FeCl 3 , 0.1 mg/L CaCl 2 and 0.013 g/L ZnCl 2 ; pH 6.0) with 10 g/L of chicken feathers and given bacterium, were incubated 48 h at 50 °C with 150 rpm.…”

“…Enzyme production of strain Oll-15 was the highest after 24 h of cultivation (14.4 U/mL), whereas strain LT-1 reached maximum production (16.6 U/mL) after 33 h. Hydrolysate from strain Oll-15 exhibited slightly better reducing activity than ascorbic acid (used as a positive control), whereas LT-1 hydrolysate did not show any significant reducing properties. Both Oll-15 and LT-1 hydrolysates showed noticeable scavenging activity of 85 and 64%, respectively, suggesting the presence of bioactive peptides in post-culture media [ 68 ].…”