Molina-Guijarro José M, Pérez Juana, Muñoz-Dorado José, Guillén Francisco, Moya Raquel, Hernández Manuel, Arias M Enriqueta
Department of Microbiology and Parasitology, University of Alcalá, Madrid, Spain.
Int Microbiol. 2009 Mar;12(1):13-21.
A newly identified extracellular laccase produced by Streptomyces ipomoea CECT 3341 (SilA) was cloned and overexpressed, and its physicochemical characteristics assessed together with its capability to decolorize and detoxify an azotype dye. Molecular analysis of the deduced sequence revealed that SilA contains a TAT-type signal peptide at the N-terminus and only two cupredoxine domains; this is consistent with reports describing two other Streptomyces laccases but contrasts with most laccases, which contain three cupredoxine domains. The heterologous expression and purification of SilA revealed that the homodimer is the only active form of the enzyme. Its stability at high pH and temperature, together with its resistance to high concentrations of NaCl and to typical laccase inhibitors such as sodium azide confirmed the unique properties of this novel laccase. The range of substrates that SilA is able to oxidize was found to be pH-dependent; at alkaline pH, SilA oxidized a wide range of phenolic compounds, including the syringyl and guayacil moieties derived from lignin. The oxidative potential of this enzyme to use phenolic compounds as natural redox mediators was shown through the coordinated action of SilA and acetosyringone (as mediator), which resulted in the complete detoxification of the azo-type dye Orange II.
克隆并过量表达了由甘薯链霉菌CECT 3341产生的一种新鉴定的细胞外漆酶（SilA），并评估了其理化特性及其对偶氮型染料的脱色和解毒能力。对推导序列的分子分析表明，SilA在N端含有一个TAT型信号肽，且仅含有两个铜蓝蛋白结构域；这与描述其他两种链霉菌漆酶的报道一致，但与大多数含有三个铜蓝蛋白结构域的漆酶形成对比。SilA的异源表达和纯化表明，同源二聚体是该酶的唯一活性形式。其在高pH和高温下的稳定性，以及对高浓度NaCl和典型漆酶抑制剂如叠氮化钠的抗性，证实了这种新型漆酶的独特性质。发现SilA能够氧化的底物范围取决于pH值；在碱性pH下，SilA氧化了多种酚类化合物，包括源自木质素的紫丁香基和愈创木酚部分。通过SilA和乙酰丁香酮（作为介质）的协同作用，展示了该酶利用酚类化合物作为天然氧化还原介质的氧化潜力，这导致偶氮型染料橙II完全解毒。
