State Key Laboratory of Animal Nutrition, College of Animal Science and Technology, China Agricultural University, Beijing, 100193, People's Republic of China.
Appl Biochem Biotechnol. 2023 Oct;195(10):6150-6167. doi: 10.1007/s12010-023-04390-4. Epub 2023 Feb 27.
The laccase gene (Lac1) was cloned from Coriolopsis trogii strain Mafic-2001. Full-length sequence of Lac1 containing 11 exons and 10 introns is composed of 2140 nucleotides (nts). mRNA of Lac1 encoded for a protein of 517 aa. Nucleotide sequence of the laccase was optimized and expressed in Pichia pastoris X-33. SDS-PAGE analysis showed that the molecular weight of the purified recombinant laccase rLac1 was about 70 kDa. The optimum temperature and pH of rLac1 were 40 ℃ and 3.0, respectively. rLac1 showed high residual activity (90%) in the solutions after 1 h incubation at the pH ranging from 2.5 to 8.0. rLac1 maintained over 60% of laccase activity at the temperatures ranging from 20 to 60 °C, and kept higher than 50% of its activity at 40 °C for 2 h. The activity of rLac1 was promoted by Cu and inhibited by Fe. Under optimal conditions, lignin degradation rates of rLac1 on the substrates of rice straw, corn stover, and palm kernel cake were 50.24%, 55.49%, and 24.43% (the lignin contents of substrates untreated with rLac1 were 100%), respectively. Treated with rLac1, the structures of agricultural residues (rice straw, corn stover, and palm kernel cake) were obviously loosened which was reflected by the analysis of scanning electron microscopy and Fourier transform infrared spectroscopy. Based on the specific activity of rLac1 on the degradation of lignin, rLac1 from Coriolopsis trogii strain Mafic-2001 has the potential for in-depth utilization of agricultural residues.
漆酶基因（Lac1）从 Coriolopsis trogii 菌株 Mafic-2001 中克隆。全长 Lac1 序列包含 11 个外显子和 10 个内含子，由 2140 个核苷酸（nts）组成。Lac1 的 mRNA 编码 517 个氨基酸的蛋白质。漆酶的核苷酸序列经过优化后在毕赤酵母 X-33 中表达。SDS-PAGE 分析表明，纯化的重组漆酶 rLac1 的分子量约为 70 kDa。rLac1 的最适温度和 pH 分别为 40℃和 3.0。rLac1 在 pH 为 2.5 至 8.0 的溶液中孵育 1 小时后，仍保持 90%的残余酶活。rLac1 在 20 至 60℃的温度范围内保持超过 60%的漆酶活性，在 40℃下保持 2 小时后，其活性仍保持在 50%以上。rLac1 的活性受 Cu 促进，受 Fe 抑制。在最佳条件下，rLac1 对水稻秸秆、玉米秸秆和棕榈仁饼底物的木质素降解率分别为 50.24%、55.49%和 24.43%（未经 rLac1 处理的底物木质素含量为 100%）。经 rLac1 处理后，农业残留物（水稻秸秆、玉米秸秆和棕榈仁饼）的结构明显疏松，这反映在扫描电子显微镜和傅里叶变换红外光谱的分析中。基于 rLac1 对木质素降解的比酶活，来自 Coriolopsis trogii 菌株 Mafic-2001 的 rLac1 具有深入利用农业残留物的潜力。
