A surfactant tolerant laccase of Meripilus giganteus
A surfactant tolerant laccase of Meripilus giganteus
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DOI：
10.1007/s11274-011-0968-z
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发表时间：
2012-04-01
影响因子：
4.1
通讯作者：
Berger, Ralf G.
中科院分区：
文献类型：
--
作者：
Schmidt, Gunnar;Krings, Ulrich;Berger, Ralf G.
从白腐菌Meripilus acuteus中分离纯化了一种漆酶（Lcc 1），其上级产率分别为34%和90%.分子排阻层析（SEC）和十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）得到的分子量为55 kDa。该酶具有3.1的等电点，并且能够在pH为2.0时氧化常见的漆酶底物2，2 '-连氮基-双（3-乙基苯并噻唑啉-6-磺酸）（ABTS），而该酶在pH 6.0时仍然能够氧化ABTS和2，6-二甲氧基苯酚（ABTS）。Lcc 1表现出低K（m）值，为8 μ M（ABTS）和80 μ M（NH3），对非酚底物ABTS的显著催化效率为37，437 k（cat）/k（m）（s（-1）mM（-1））。该漆酶对高浓度的金属离子、EDTA和表面活性剂具有很高的稳定性，具有很大的生物技术潜力。此外，Lcc 1表现出增加的活性以及在表面活性剂存在下的稳定性的显著提高。从肽片段推导简并引物。lcc 1的完整编码序列被确定为1，551 bp，并通过扩增包括12个内含子的2，214 bp基因组序列来确认。lcc 1基因的516个氨基酸序列与微孔刚孔菌漆酶的同源性为82%，相似性为90%。序列数据可以帮助理论研究和酶工程的努力，以创建漆酶与金属离子和双极性化合物的稳定性提高。
A laccase (Lcc1) from the white-rot fungus Meripilus giganteus was purified with superior yields of 34% and 90% by conventional chromatography or by foam separation, respectively. Size exclusion chromatography (SEC) and sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) yielded a molecular mass of 55 kDa. The enzyme possessed an isoelectric point of 3.1 and was able to oxidize the common laccase substrate 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) at a pH of 2.0, whereas the enzyme was still able to oxidize ABTS and 2,6-dimethoxyphenol (DMP) at pH 6.0. Lcc1 exhibited low K (m) values of 8 mu M (ABTS) and 80 mu M (DMP) and remarkable catalytic efficiency towards the non-phenolic substrate ABTS of 37,437 k (cat)/k (m) (s(-1) mM(-1)). The laccase showed a high stability towards high concentrations of various metal ions, EDTA and surfactants indicating a considerable biotechnological potential. Furthermore, Lcc1 exhibited an increased activity as well as a striking boost of stability in the presence of surfactants. Degenerated primers were deduced from peptide fragments. The complete coding sequence of lcc1 was determined to 1,551 bp and confirmed via amplification of the 2,214 bp genomic sequence which included 12 introns. The deduced 516 amino acid (aa) sequence of the lcc1 gene shared 82% identity and 90% similarity with a laccase from Rigidoporus microporus. The sequence data may aid theoretical studies and enzyme engineering efforts to create laccases with an improved stability towards metal ions and bipolar compounds.