Larsbrink Johan, Tuveng Tina R, Pope Phillip B, Bulone Vincent, Eijsink Vincent G H, Brumer Harry, McKee Lauren S
Norwegian University of Life Sciences, NMBU, Department of Chemistry, Biotechnology and Food Sciences, PO Box 5003, NO-1432 Ås, Norway; Wallenberg Wood Science Center, Division of Industrial Biotechnology, Department of Biology and Biological Engineering, Chalmers University, 412 96 Gothenburg, Sweden.
Norwegian University of Life Sciences, NMBU, Department of Chemistry, Biotechnology and Food Sciences, PO Box 5003, NO-1432 Ås, Norway.
J Proteomics. 2017 Mar 6;156:63-74. doi: 10.1016/j.jprot.2017.01.003. Epub 2017 Jan 6.
Together with fungi, saprophytic bacteria are central to the decomposition and recycling of biomass in forest environments. The Bacteroidetes phylum is abundant in diverse habitats, and several species have been shown to be able to deconstruct a wide variety of complex carbohydrates. The genus Chitinophaga is often enriched in hotspots of plant and microbial biomass degradation. We present a proteomic assessment of the ability of Chitinophaga pinensis to grow on and degrade mannan polysaccharides, using an agarose plate-based method of protein collection to minimise contamination with exopolysaccharides and proteins from lysed cells, and to reflect the realistic setting of growth on a solid surface. We show that select Polysaccharide Utilisation Loci (PULs) are expressed in different growth conditions, and identify enzymes that may be involved in mannan degradation. By comparing proteomic and enzymatic profiles, we show evidence for the induced expression of enzymes and PULs in cells grown on mannan polysaccharides compared with cells grown on glucose. In addition, we show that the secretion of putative biomass-degrading enzymes during growth on glucose comprises a system for nutrient scavenging, which employs constitutively produced enzymes.
Chitinophaga pinensis belongs to a bacterial genus which is prominent in microbial communities in agricultural and forest environments, where plant and fungal biomass is intensively degraded. Such degradation is hugely significant in the recycling of carbon in the natural environment, and the enzymes responsible are of biotechnological relevance in emerging technologies involving the deconstruction of plant cell wall material. The bacterium has a comparatively large genome, which includes many uncharacterised carbohydrate-active enzymes. We present the first proteomic assessment of the biomass-degrading machinery of this species, focusing on mannan, an abundant plant cell wall hemicellulose. Our findings include the identification of several novel enzymes, which are promising targets for future biochemical characterisation. In addition, the data indicate the expression of specific Polysaccharide Utilisation Loci, induced in the presence of different growth substrates. We also highlight how a constitutive secretion of enzymes which deconstruct microbial biomass likely forms part of a nutrient scavenging process.
腐生细菌与真菌一起，在森林环境中生物质的分解和循环利用中起着核心作用。拟杆菌门在多种生境中数量丰富，并且已证明有几个物种能够解构多种复杂碳水化合物。几丁质噬菌属通常在植物和微生物生物质降解的热点区域富集。我们采用基于琼脂糖平板的蛋白质收集方法，对松树几丁质噬菌体能在甘露聚糖多糖上生长和降解的能力进行了蛋白质组学评估，以尽量减少胞外多糖和裂解细胞中蛋白质的污染，并反映在固体表面生长的实际情况。我们表明，特定的多糖利用位点（PULs）在不同的生长条件下表达，并鉴定出可能参与甘露聚糖降解的酶。通过比较蛋白质组学和酶学谱，我们证明了与在葡萄糖上生长的细胞相比，在甘露聚糖多糖上生长的细胞中酶和PULs的诱导表达。此外，我们表明，在葡萄糖上生长期间推定的生物质降解酶的分泌包括一个营养清除系统，该系统使用组成型产生的酶。
松树几丁质噬菌属属于一个在农业和森林环境的微生物群落中占主导地位的细菌属，在这些环境中植物和真菌生物质被大量降解。这种降解在自然环境中的碳循环中具有极其重要的意义，并且所涉及的酶在涉及解构植物细胞壁材料的新兴技术中具有生物技术相关性。该细菌具有相对较大的基因组，其中包括许多未表征的碳水化合物活性酶。我们首次对该物种的生物质降解机制进行了蛋白质组学评估，重点是甘露聚糖，一种丰富的植物细胞壁半纤维素。我们的发现包括鉴定出几种新型酶，它们是未来生化表征的有希望的目标。此外，数据表明在不同生长底物存在下诱导表达特定的多糖利用位点。我们还强调了解构微生物生物质的酶的组成型分泌可能如何形成营养清除过程的一部分。
