Bioprocess Engineering Group, Institute of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
Sci Rep. 2021 Feb 25;11(1):4573. doi: 10.1038/s41598-021-84267-w.
Thermostability and substrate specificity of proteases are major factors in their industrial applications. rEla is a novel recombinant cysteine protease obtained from a thermophilic bacterium, Cohnella sp.A01 (PTCC No: 1921). Herein, we were interested in recombinant production and characterization of the enzyme and finding the novel features in comparison with other well-studied cysteine proteases. The bioinformatics analysis showed that rEla is allosteric cysteine protease from DJ-1/ThiJ/PfpI superfamily. The enzyme was heterologously expressed and characterized and the recombinant enzyme molecular mass was 19.38 kD which seems to be smaller than most of the cysteine proteases. rEla exhibited acceptable activity in broad pH and temperature ranges. The optimum activity was observed at 50℃ and pH 8 and the enzyme showed remarkable stability by keeping 50% of residual activity after 100 days storage at room temperature. The enzyme K and V values were 21.93 mM, 8 U/ml, respectively. To the best of our knowledge, in comparison with the other characterized cysteine proteases, rEla is the only reported cysteine protease with collagen specificity. The enzymes activity increases up to 1.4 times in the presence of calcium ion (2 mM) suggesting it as the enzyme's co-factor. When exposed to surfactants including Tween20, Tween80, Triton X-100 and SDS (1% and 4% v/v) the enzyme activity surprisingly increased up to 5 times.
蛋白酶的热稳定性和底物特异性是其工业应用的主要因素。rEla 是一种新型的重组半胱氨酸蛋白酶，从嗜热细菌 Cohnella sp.A01（PTCC No：1921）中获得。在此，我们对该酶的重组生产和特性感兴趣，并与其他研究充分的半胱氨酸蛋白酶进行比较，寻找新的特征。生物信息学分析表明，rEla 是 DJ-1/ThiJ/PfpI 超家族的变构半胱氨酸蛋白酶。该酶进行了异源表达和特性分析，重组酶的分子量为 19.38 kD，似乎比大多数半胱氨酸蛋白酶小。rEla 在广泛的 pH 和温度范围内表现出可接受的活性。最佳活性在 50℃和 pH 8 下观察到，该酶在室温下储存 100 天后保持 50%的残余活性，显示出显著的稳定性。酶的 K 和 V 值分别为 21.93 mM 和 8 U/ml。据我们所知，与其他表征的半胱氨酸蛋白酶相比，rEla 是唯一报道的具有胶原蛋白特异性的半胱氨酸蛋白酶。在钙离子（2 mM）存在下，酶的活性增加了 1.4 倍，表明其为酶的辅助因子。当暴露于表面活性剂（包括吐温 20、吐温 80、Triton X-100 和 SDS（1%和 4%v/v）时，酶的活性令人惊讶地增加了 5 倍。
