Sun Jiayi, Ren Jun, Xu Li, Jia Lingyun
Department of Life Science and Biotechnology, Dalian University of Technology, Dalian 116024, China.
Se Pu. 2010 Sep;28(9):872-6.
Monoamine oxidase B (MAOB) was purified from porcine liver by solubilization with lysis buffer containing 1% Triton X-100, precipitation with 20%-50% ammonium sulfate, isolation with hydrophobic chromatography and anion exchange chromatography. The purification fold was 18.2. The specific activity was 135 U/mg. The purified enzyme appeared homogeneous by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and it had a relative molecular mass of about 60,000. The identification of the enzyme was confirmed by high performance liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). As MAOB is a membrane enzyme, a key step to the successful purification was the use of Phenyl-Sepharose CL-4B with phenyl density of 75.7 micromol/mL. The results showed that this approach could effectively isolate MAOB from porcine liver to yield an enzyme with high purity and specific activity.
单胺氧化酶B（MAOB）通过以下步骤从猪肝中纯化：用含1% Triton X-100的裂解缓冲液溶解，用20%-50%硫酸铵沉淀，通过疏水色谱和阴离子交换色谱分离。纯化倍数为18.2。比活性为135 U/mg。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE），纯化后的酶呈现均一性，其相对分子质量约为60,000。通过高效液相色谱-电喷雾电离串联质谱（HPLC-ESI-MS/MS）确认了该酶的身份。由于MAOB是一种膜酶，成功纯化的关键步骤是使用苯基密度为75.7微摩尔/毫升的苯基琼脂糖凝胶CL-4B。结果表明，该方法可以有效地从猪肝中分离出MAOB，得到具有高纯度和比活性的酶。
