PDF(Sci-hub)
Abstract：
A white rot fungus Cerrena unicolor has been identified as an important source of laccase, unfortunately regulation of this enzyme genes expression is poorly understood. Using 1D and 2D PAGE and LC-MS/MS, laccase isoenzymes were investigated in the liquid filtrate of C. unicolor culture. The level of expression of laccase genes was measured using qPCR. The elevated concentrations of copper and manganese in the medium caused greatest change in genes expression and three laccase transcripts were significantly affected after culture temperature was decreased from 28 to 4 °C or increased to 40 °C. The small differences in the PAGE band intensities of individual laccase proteins were also observed, indicating that given compound affect particular laccase\'s transcript. Analyses of laccase-specific activity, at all tested conditions, showed the increased activities as compared to the control, suggesting that enzyme is regulated at the post-translational stage. We observed that the aspartic protease purified from C. unicolor, significantly stimulate laccase activity. Moreover, electrochemical analysis of protease-treated laccase sample had 5 times higher redox peaks. The obtained results indicate that laccases released by C. unicolor are regulated at transcriptional, translational, and at the post-translational steps of gene expression helping fungus adapt to the environmental changes.
摘要：
白腐真菌Cerrenaunicolor已被确定为漆酶的重要来源，不幸的是，人们对这种酶基因表达的调节知之甚少。使用1D和2DPAGE和LC-MS/MS，在C.单色培养物的液体滤液中研究漆酶同工酶。使用qPCR测量漆酶基因的表达水平。培养基中铜和锰浓度的升高导致基因表达的最大变化，培养温度从28℃降低到4℃或升高到40℃后，三种漆酶转录物受到显着影响。还观察到单个漆酶蛋白的PAGE条带强度的小差异，表明给定的化合物会影响特定的漆酶的转录本。漆酶特异性活性的分析，在所有测试条件下，显示与对照组相比活动增加，这表明酶在翻译后阶段受到调节。我们观察到从C.unicolor中纯化的天冬氨酸蛋白酶，显著刺激漆酶活性。此外，电化学分析蛋白酶处理的漆酶样品具有高5倍的氧化还原峰。获得的结果表明，由C.unicolor释放的漆酶在转录受到调控，翻译,在基因表达的翻译后步骤中帮助真菌适应环境变化。
