Currie D H, Guss A M, Herring C D, Giannone R J, Johnson C M, Lankford P K, Brown S D, Hettich R L, Lynd L R
Thayer School of Engineering, Dartmouth College, Hanover, New Hampshire, USA Mascoma Corporation, Lebanon, New Hampshire, USA.
Thayer School of Engineering, Dartmouth College, Hanover, New Hampshire, USA Oak Ridge National Laboratory, Oak Ridge, Tennessee, USA.
Appl Environ Microbiol. 2014 Aug;80(16):5001-11. doi: 10.1128/AEM.00998-14. Epub 2014 Jun 6.
Thermoanaerobacterium saccharolyticum, a Gram-positive thermophilic anaerobic bacterium, grows robustly on insoluble hemicellulose, which requires a specialized suite of secreted and transmembrane proteins. We report here the characterization of proteins secreted by this organism. Cultures were grown on hemicellulose, glucose, xylose, starch, and xylan in pH-controlled bioreactors, and samples were analyzed via spotted microarrays and liquid chromatography-mass spectrometry. Key hydrolases and transporters employed by T. saccharolyticum for growth on hemicellulose were, for the most part, hitherto uncharacterized and existed in two clusters (Tsac_1445 through Tsac_1464 for xylan/xylose and Tsac_1344 through Tsac_1349 for starch). A phosphotransferase system subunit, Tsac_0032, also appeared to be exclusive to growth on glucose. Previously identified hydrolases that showed strong conditional expression changes included XynA (Tsac_1459), XynC (Tsac_0897), and a pullulanase, Apu (Tsac_1342). An omnipresent transcript and protein making up a large percentage of the overall secretome, Tsac_0361, was tentatively identified as the primary S-layer component in T. saccharolyticum, and deletion of the Tsac_0361 gene resulted in gross morphological changes to the cells. The view of hemicellulose degradation revealed here will be enabling for metabolic engineering efforts in biofuel-producing organisms that degrade cellulose well but lack the ability to catabolize C5 sugars.
嗜糖栖热厌氧杆菌是一种革兰氏阳性嗜热厌氧菌，能在不溶性半纤维素上旺盛生长，这需要一套特殊的分泌蛋白和跨膜蛋白。我们在此报告该生物体分泌蛋白的特性。在pH受控的生物反应器中，将培养物置于半纤维素、葡萄糖、木糖、淀粉和木聚糖上培养，通过点阵微阵列和液相色谱 - 质谱联用对样品进行分析。嗜糖栖热厌氧杆菌在半纤维素上生长所使用的关键水解酶和转运蛋白，在很大程度上此前未被表征，且存在于两个簇中（木聚糖/木糖相关的从Tsac_1445到Tsac_1464，淀粉相关的从Tsac_1344到Tsac_1349）。一种磷酸转移酶系统亚基Tsac_0032似乎也仅在葡萄糖上生长时才存在。先前鉴定出的显示出强烈条件性表达变化的水解酶包括木聚糖酶A（Tsac_1459）、木聚糖酶C（Tsac_0897）和一种支链淀粉酶Apu（Tsac_1342）。一种普遍存在的转录本和蛋白Tsac_0361在整个分泌组中占很大比例，初步鉴定为嗜糖栖热厌氧杆菌的主要S层成分，删除Tsac_0361基因导致细胞出现明显的形态变化。此处揭示的半纤维素降解观点将有助于在生物燃料生产生物体中进行代谢工程研究，这些生物体能够很好地降解纤维素，但缺乏分解C5糖的能力。
