Prajapati Vimal, Patel Honey, Trivedi Ujjval, Patel Kamlesh
B R D School of Biosciences, Sardar Patel University, Sardar Patel Maidan, Vadtal Road, Vallabh Vidyanagar, Gujarat, India.
J Basic Microbiol. 2014 Sep;54(9):976-83. doi: 10.1002/jobm.201300065. Epub 2013 May 26.
Lipase of Cellulomonas flavigena UNP3 was purified by two-step purification process comprising ammonium sulfate precipitation followed by gel permeation chromatography (GPC). The recovery of lipase after GPC was found to be 1.70% with 20.98-fold increase in specific activity. The molecular weight of lipase protein was found to be 45.2 kDa by SDS-PAGE. Activation energy for p-nitrophenol palmitate (pNPP) hydrolysis was 26.45 kJ mol(-1) , while temperature quotient (Q10 ) was found to be 1.64. The enzyme was found to be stable over wide pH range and thermally stable at 30-40 °C up to 60 min of incubation while exhibited maximum activity at 30 °C with pH 7.0. Vmax , Km , and Kcat for pNPP were found to be 666.71 U ml(-1) , 1.33 mM (pNPP) and 433 min(-1) , respectively. Activation energy for irreversible inactivation Ea(d) of lipase was 64.32 kJ mol(-1) . Thermodynamic parameters of irreversible inactivation of lipase and pNPP hydrolysis were also determined.
黄褐纤维单胞菌UNP3脂肪酶通过两步纯化过程进行纯化，该过程包括硫酸铵沉淀，随后进行凝胶渗透色谱（GPC）。GPC后脂肪酶的回收率为1.70%，比活增加了20.98倍。通过SDS-PAGE测定脂肪酶蛋白的分子量为45.2 kDa。对硝基苯酚棕榈酸酯（pNPP）水解的活化能为26.45 kJ mol(-1)，而温度系数（Q10）为1.64。发现该酶在较宽的pH范围内稳定，在30 - 40°C下热稳定，孵育60分钟，同时在30°C、pH 7.0时表现出最大活性。pNPP的Vmax、Km和Kcat分别为666.71 U ml(-1)、1.33 mM（pNPP）和433 min(-1)。脂肪酶不可逆失活的活化能Ea(d)为64.32 kJ mol(-1)。还测定了脂肪酶不可逆失活和pNPP水解的热力学参数。
