In vitro fibrinolytic activity of an enzyme purified from Bacillus amyloliquefaciens strain KJ10 isolated from soybean paste

Abstract

A fibrinolytic protease secreting producing Bacillus amyloliquefaciens strain KJ10 was initially screened from the fermented soybean. Maximum productivity was obtained in the culture medium after 40 h incubation, 34 °C incubation temperature at pH 8.0. Fibrinolytic protease production was enhanced in the culture medium with 1% sucrose (3712 ± 52 U/mL), 1% (w/v) yeast extract (3940 ± 28 U/mL) and 0.1% MgSO 4 (3687 ± 38 U/mL). Enzyme was purified up to 22.9-fold with 26%recovery after Q-Sepharose HP column chromatography. After three steps purification, enzyme activity was 1606U/mg and SDS-PAGE analysis revealed 29 kDa protein and enzyme band was detected by zymograpy. Enzyme was highly active at pH 8.0, at wide temperature ranges (40 °C − 55 °C) and was activated by Mn 2+ (102 ± 3.1%) and Mg 2+ (101.4 ± 2.9%) ions. The purified fibrinolytic enzyme was highly specific against N-Suc-Ala-Ala-Pro-Phe- p NA (189 mmol/min/mL) and clot lytic activity reached 28 ± 1.8% within 60 min in vitro . The purified fibrinolytic enzyme showed least erythrocytic lysis activity confirmed safety to prevent various health risks, including hemolytic anemia. Based on this study, administration of fibrinolytic enzyme from B. amyloliquefaciens strain KJ10 is safe for clinical applications.

Novel Fibrinolytic Enzyme by Scopulariopsis brevicaulis OS 3456: Production, Characterization, In vitro, and In vivo Activity

“…Furthermore, the enzyme became unstable at temperatures above 50 o C, verifying that the enzyme has somewhat good thermal stability. These findings are the same as those of fibrinolytic enzymes from Streptomyces radiopugnans VITSD8 (Dhamodharan et al, 2019) and Bacillus amyloliquefaciens strain KJ10 (Rajaselvam et al, 2021). Meanwhile, the fibrinolytic enzyme produced by Biji et al (2016) was thermophilic with an optimum temperature of 50 o C, and the enzyme obtained by Kornienko et al (2021) was alkaline with a pH of 10.0.…”

Microbial Fibrinolytic Enzymes as Anti-Thrombotics: Production, Characterisation and Prodigious Biopharmaceutical Applications

“…Such methodologies are used either individually or in combinations, accompanied by chromatographic techniques for further purification. Chromatofocusing, fast protein liquid chromatography, high performance liquid chromatography, affinity column chromatography, gel filtration chromatography, ion exchange chromatography and hydrophobic interaction chromatography are commonly employed techniques for fibrinolytic enzyme purification [2,5,[100][101][102][103][104][105][106][107][108]110,[162][163][164][165][166][167][168]. Some recent purification studies of microbial fibrinolytic enzymes employed by researchers are discussed below.…”

Thrombolytic Enzymes of Microbial Origin: A Review

“…There are several known sources of fibrinolytic enzymes, including plants, animals, and microbes. Among them, microbial-derived fibrinolytic enzymes have gained particular interest because they possess the following advantages: (i) cost-effective production, (ii) fewer to no side effects [ 6 ], (iii) broad biochemical diversity, (iv) mass-culture feasibility, and (v) allow genetic manipulation. Hence, various microorganisms have been used for the isolation of fibrinolytic enzymes, including bacteria, fungi, and algae.…”