Centre for Novel Agricultural Products, Department of Biology, University of York, York, YO10 5DD, UK.
Bioscience Technology Facility, Department of Biology, University of York, York, YO10 5DD, UK.
Sci Rep. 2017 May 24;7(1):2356. doi: 10.1038/s41598-017-02506-5.
Microbial communities metabolize plant biomass using secreted enzymes; however, identifying extracellular proteins tightly bound to insoluble lignocellulose in these microbiomes presents a challenge, as the rigorous extraction required to elute these proteins also lyses the microbes associated with the plant biomass releasing intracellular proteins that contaminate the metasecretome. Here we describe a technique for targeting the extracellular proteome, which was used to compare the metasecretome and meta-surface-proteome of two lignocellulose-degrading communities grown on wheat straw and rice straw. A combination of mass spectrometry-based proteomics coupled with metatranscriptomics enabled the identification of a unique secretome pool from these lignocellulose-degrading communities. This method enabled us to efficiently discriminate the extracellular proteins from the intracellular proteins by improving detection of actively secreted and transmembrane proteins. In addition to the expected carbohydrate active enzymes, our new method reveals a large number of unknown proteins, supporting the notion that there are major gaps in our understanding of how microbial communities degrade lignocellulosic substrates.
微生物群落利用分泌的酶代谢植物生物质；然而，在这些微生物群落中，鉴定紧密结合于不溶性木质纤维素的细胞外蛋白质是一项挑战，因为为了洗脱这些蛋白质所需的严格提取也会裂解与植物生物质相关的微生物，从而释放出污染代谢组的细胞内蛋白质。在这里，我们描述了一种靶向细胞外蛋白质组的技术，该技术用于比较在小麦秸秆和水稻秸秆上生长的两个木质纤维素降解群落的代谢组和元表面蛋白质组。基于质谱的蛋白质组学与元转录组学的结合，使我们能够从这些木质纤维素降解群落中鉴定出一个独特的分泌组池。这种方法通过提高对活跃分泌和跨膜蛋白质的检测，使我们能够有效地将细胞外蛋白质与细胞内蛋白质区分开来。除了预期的碳水化合物活性酶外，我们的新方法还揭示了大量未知蛋白质，这支持了这样一种观点，即我们对微生物群落如何降解木质纤维素底物的理解存在重大差距。
