College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, China.
Bioresour Technol. 2011 Feb;102(3):3126-37. doi: 10.1016/j.biortech.2010.10.079. Epub 2010 Oct 23.
The laccase gene lac48424-1 and its corresponding full-length cDNA were cloned and characterized from a novel white-rot fungi Trametes sp. 48424 which had the high yield of laccase and strong ability for decolorizing different dyes. The 1563 bp full-length cDNA of lac48424-1 encoded a mature laccase protein containing 499 amino acids preceded by a signal peptide of 21 amino acids. The deduced protein sequence of LAC48424-1 showed high similarity with other known fungal laccases and contained four copper-binding conserved domains of typical laccase protein. The functionality of lac48424-1 gene encoding active laccase was verified by expressing the gene in the yeast Pichia pastoris successfully. It was found that the recombinant laccase produced by the yeast transformant could decolorize different dyes. The 5'-flanking sequence upstream of start codon was obtained by Self-Formed Adaptor PCR. Many putative cis-acting responsive elements involved in the transcriptional regulation were identified in the promoter region of lac48424-1.
从一株产漆酶量高且对不同染料具有较强脱色能力的新型白腐真菌 Trametes sp. 48424 中克隆并鉴定了漆酶基因 lac48424-1 及其全长 cDNA。lac48424-1 的全长 cDNA 为 1563bp，编码一个成熟的漆酶蛋白，该蛋白含有 499 个氨基酸，前端有 21 个氨基酸的信号肽。LAC48424-1 推导的蛋白序列与其他已知真菌漆酶具有高度相似性，并且包含典型漆酶蛋白的四个铜结合保守结构域。通过成功在毕赤酵母 Pichia pastoris 中表达该基因来验证编码活性漆酶的 lac48424-1 基因的功能。结果发现，酵母转化子产生的重组漆酶可以使不同的染料脱色。通过自我形成接头 PCR 获得了起始密码子上游的 5'侧翼序列。在 lac48424-1 的启动子区域中鉴定到了许多参与转录调控的顺式作用响应元件。
