Biundo Antonino, Hromic Altijana, Pavkov-Keller Tea, Gruber Karl, Quartinello Felice, Haernvall Karolina, Perz Veronika, Arrell Miriam S, Zinn Manfred, Ribitsch Doris, Guebitz Georg M
Institute of Environmental Biotechnology, University of Natural Resources and Life Science (BOKU), Konrad Lorenz Strasse 22, 3430, Tulln an der Donau, Austria.
Austrian Centre for Industrial Biotechnology (ACIB), Petersgasse 14, 8010, Graz, Austria.
Appl Microbiol Biotechnol. 2016 Feb;100(4):1753-1764. doi: 10.1007/s00253-015-7031-1.
Certain α/β hydrolases have the ability to hydrolyze synthetic polyesters. While their partial hydrolysis has a potential for surface functionalization, complete hydrolysis allows recycling of valuable building blocks. Although knowledge about biodegradation of these materials is important regarding their fate in the environment, it is currently limited to aerobic organisms. A lipase from the anaerobic groundwater organism Pelosinus fermentans DSM 17108(PfL1) was cloned and expressed in Escherichia coli BL21-Gold (DE3) and purified from the cell extract. Biochemical characterization with small substrates showed thermoalkalophilic properties (Topt=50 °C, pHopt=7.5) and higher activity towards para-nitrophenyl octanoate (12.7 U mg(-1)) compared to longer and shorter chain lengths (C14 0.7 U mg(-1) and C2 4.3 U mg(-1), respectively). Crystallization and determination of the 3-D structure displayed the presence of a lid structure and a zinc ion surrounded by an extra domain. These properties classify the enzyme into the I.5 lipase family. PfL1 is able to hydrolyze poly(1,4-butylene adipate-co-terephthalate) (PBAT) polymeric substrates. The hydrolysis of PBAT showed the release of small building blocks as detected by liquid chromatography mass spectrometry (LC-MS). Protein dynamics seem to be involved with lid opening for the hydrolysis of PBAT by PfL1.
某些α/β水解酶具有水解合成聚酯的能力。虽然它们的部分水解具有表面功能化的潜力，但完全水解可实现有价值的结构单元的回收利用。尽管关于这些材料在环境中的生物降解知识对于了解它们的归宿很重要，但目前仅限于需氧生物。从厌氧地下水微生物费氏泥杆菌DSM 17108（PfL1）中克隆出一种脂肪酶，并在大肠杆菌BL21-Gold（DE3）中表达，然后从细胞提取物中纯化出来。用小分子底物进行的生化特性分析显示出嗜热嗜碱特性（最适温度=50℃，最适pH=7.5），与较长和较短链长的底物（分别为C14 0.7 U mg-1和C2 4.3 U mg-1）相比，对对硝基苯基辛酸酯的活性更高（12.7 U mg-1）。晶体结构的结晶和三维结构测定显示存在一个盖子结构和一个被额外结构域包围的锌离子。这些特性将该酶归类为I.5脂肪酶家族。PfL1能够水解聚（1,4-丁二醇己二酸酯-对苯二甲酸酯）（PBAT）聚合物底物。通过液相色谱-质谱联用（LC-MS）检测发现，PBAT的水解显示出小分子结构单元的释放。蛋白质动力学似乎与PfL1水解PBAT时盖子的打开有关。
