Abstract

Alkaline proteases having activity and stability at alkaline pH possess a large variety of applications in many industries. Growing renewed interest urges the need to find a single alkaline protease with promising properties to be used in different industrial processes. Herein, alkaline proteases produced through fermentation of cheap and easily available organic municipal solid wastes by Bacillus subtilis AKAL7 and Exiguobacterium indicum AKAL11 were purified to investigate their kinetic and thermodynamic parameters, detergent compatibility, dehairing and feather-degrading capability. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed that the purified protease from B. subtilis and E. indicum had molecular mass of ∼45 and 75 kDa, respectively. The protease from B. subtilis and E. indicum showed highest activity at 55 and 50 °C having low K _m 1.17 and 0.567 mg/mL and high V _max 416.67 and 333.33 µmole/min, respectively. The activation energy and temperature quotient of protease from B. subtilis and E. indicum were 26.52 and 65.75 kJ/mole, and 1.0004 and 1.0003 at 20–55 and 20–50 °C, respectively. Thermodynamics analysis revealed the formation of more ordered enzyme–substrate complexes along with spontenity of enzyme reaction. The protease from E. indicum exhibited better compatibility at higher concentration of detergents compared to that from B. subtilis. However, both proteases could retain more than 80% of the activity in the presence of 0.1% commercial laundry detergents. The purified protease from the both sources could degrade almost 90% of barbs and 40% of dry weight of the native feather and that from E. indicum could dehair cow skin. Results reported herein suggest that the alkaline protease from B. subtilis AKAL7 and E. indicum AKAL11 has biotechnological implications in detergent, leather and poultry feather processing industries.

摘要

在碱性pH下具有活性和稳定性的碱性蛋白酶在许多工业中具有多种应用。越来越多的新兴趣促使人们迫切需要找到一种具有前途特性的单一碱性蛋白酶，以用于不同的工业过程。本文中，对枯草芽孢杆菌AKAL7和靛蓝杆菌（ Exiguobacterium indicum） AKAL11发酵廉价易得的有机城市固体废物产生的碱性蛋白酶进行了纯化，以研究其动力学和热力学参数，去污剂相容性，脱毛和降解羽毛的能力。十二烷基硫酸钠聚丙烯酰胺凝胶电泳显示，枯草芽孢杆菌和印度大肠埃希菌纯化的蛋白酶分子质量分别约为45和75 kDa。枯草芽孢杆菌和印度大肠杆菌的蛋白酶在55和50°C下显示最高活性，分别具有低K _m 1.17和0.567 mg / mL和高V _max 416.67和333.33 µmole / min。在20–55和20–50°C下，枯草芽孢杆菌和印度小肠杆菌的蛋白酶的活化能和温度商分别为26.52和65.75 kJ / mol，以及1.0004和1.0003。热力学分析表明，随着酶反应的自发性，形成了更多有序的酶-底物复合物。来自印度大肠埃希菌的蛋白酶与枯草芽孢杆菌相比，在较高浓度的洗涤剂中具有更好的相容性。但是，在0.1％的商用洗衣剂存在下，两种蛋白酶都可以保留80％以上的活性。来自两种来源的纯化的蛋白酶可降解近90％的倒钩和40％的天然羽毛干重，而来自印度肠杆菌的可降解毛发。本文报道的结果表明，来自枯草芽孢杆菌AKAL7和靛蓝大肠杆菌AKAL11的碱性蛋白酶在洗涤剂，皮革和家禽羽毛加工工业中具有生物技术意义。