Screening of Elastase, Fibrinogenase and Collagenase Among Bacillus Strains Isolated from Pine Seeds

Abstract

The dominant position on the market of industrial enzymes (over 60%) is occupied by proteases, two-thirds of which are of microbial origin. Of particular value are stable proteases with specificity for elastin, fibrinogen, and collagen. The search for new producers among different econiches is an urgent task for obtaining enzymes with unique properties. Endophytic bacteria, in particular, representatives of the genus Bacillus, which colonize the internal tissues of plants, are a promising but little-studied source of bioactive compounds. Since data on the proteolytic potential of endophytes of Scots pine seeds (Pinus sylvestris L.) in the available literature are fragmentary, the aim of the work was to screen bacterial isolates from this econiche for the presence of enzymes with elastase, fibrinogenase, and collagenase activity. Methods. The objects of the study were 13 cultures isolated from Scots pine seeds. The bacteria were grown in submerged cultivation at 28 °C, with a stirring speed of the nutrient medium of 230 rpm for 3 days. Methods for determining proteolytic (elastolytic, fibrinogenolytic, and collagenolytic) activity in the supernatant of the culture liquid were used. Results. The study of the ability of the supernatants of the studied cultures to hydrolyze various proteolytic substrates showed that the strains Bacillus sp. E12 and E5 were promising for further research. In their complex enzyme preparations, elastase activity was manifested in the same pH range from 6 to 12, with a pH optimum of 11 and 8 and temperatures from 4 to 70 °C for Bacillus sp. E12 and E5, respectively. However, the thermooptima of elastase activity in the preparations of both strains were slightly different and were 4 and 50 °C for Bacillus sp. E12 and E5, respectively. As for fibrinogenolytic activity, the enzyme preparations of Bacillus sp. E12 and E5 differed significantly in activity (6.8 and 1.05 U/mL) and were characterized by almost the same pH optimum (10 and 9) but differed sharply in thermooptima (38 and 60 °C). Conclusions. For the first time, effective producers of proteases with elastase and fibrinogenolytic activity have been identified from bacteria isolated from Scots pine seeds. The results obtained expand understanding of the biological activity of endophytic bacteria, inhabitants of Scots pine.