Bhosale Hemlata, Shaheen Uzma, Kadam Tukaram
DST-FIST Sponsored School of Life Sciences, Swami Ramanand Teerth Marathwada University, Nanded 431606, India.
Enzyme Res. 2016;2016:4170684. doi: 10.1155/2016/4170684. Epub 2016 Jan 21.
Hyperthermostable alkaline lipase from Bacillus sonorensis 4R was purified and characterized. The enzyme production was carried out at 80°C and 9.0 pH in glucose-tween inorganic salt broth under static conditions for 96 h. Lipase was purified by anion exchange chromatography by 12.15 fold with a yield of 1.98%. The molecular weight of lipase was found to be 21.87 KDa by SDS-PAGE. The enzyme activity was optimal at 80°C with t 1/2 of 150 min and at 90°C, 100°C, 110°C, and 120°C; the respective values were 121.59 min, 90.01 min, 70.01 min, and 50 min. The enzyme was highly activated by Mg and t 1/2 values at 80°C were increased from 150 min to 180 min when magnesium and mannitol were added in combination. The activation energy calculated from Arrhenius plot was 31.102 KJ/mol. At 80-120°C, values of ΔH and ΔG were in the range of 28.16-27.83 KJ/mol and 102.79 KJ/mol to 111.66 KJ/mol, respectively. Lipase activity was highest at 9.0 pH and stable for 2 hours at this pH at 80°C. Pretreatment of lipase with MgSO4 and CaSO4 stimulated enzyme activity by 249.94% and 30.2%, respectively. The enzyme activity was greatly reduced by CoCl2, CdCl2, HgCl2, CuCl2, Pb(NO3)2, PMSF, orlistat, oleic acid, iodine, EDTA, and urea.
对来自索诺拉芽孢杆菌4R的超嗜热碱性脂肪酶进行了纯化和表征。在静态条件下，于葡萄糖 - 吐温无机盐肉汤中，80°C和pH 9.0下进行96小时的酶生产。通过阴离子交换色谱法对脂肪酶进行纯化，纯化倍数为12.15倍，产率为1.98%。通过SDS - PAGE测定，脂肪酶的分子量为21.87 kDa。该酶在80°C时活性最佳，半衰期为150分钟；在90°C、100°C、110°C和120°C时，相应的值分别为121.59分钟、90.01分钟、70.01分钟和50分钟。该酶被Mg高度激活，当同时添加镁和甘露醇时，80°C下的半衰期从150分钟增加到180分钟。根据阿伦尼乌斯图计算出的活化能为31.102 KJ/mol。在80 - 120°C下，ΔH和ΔG的值分别在28.16 - 27.83 KJ/mol和102.79 KJ/mol至111.66 KJ/mol范围内。脂肪酶活性在pH 9.0时最高，在80°C下此pH值下稳定2小时。用MgSO4和CaSO4预处理脂肪酶分别使酶活性提高了249.94%和30.2%。CoCl2、CdCl2、HgCl2、CuCl2、Pb(NO3)2、PMSF、奥利司他、油酸、碘、EDTA和尿素会大大降低该酶的活性。
