Genomic analysis, identification and characterization of lignin degradation pathway and biofuel production by the Gram-positive bacterium Paenibacillus polymyxa CR1.

从木质纤维素生物质衍生的生物燃料有望成为清洁和可再生能源和化学品的替代来源。然而，脱木质素或木质素分解是生物燃料和生物炼制的瓶颈。因此，了解细菌中木质素降解途径、生物燃料生产及其调控机制具有重要意义。作为我的研究计划的一部分，旨在开发新的战略，可持续农业和环境，我的实验室最近的特点Paenibacilluspolymyxa CR 1，革兰氏阳性，孢子形成，兼性厌氧菌，我们从降解玉米根分离在南部安大略。我们发现P. polymyxa CR 1具有很强的降解和利用木质素、纤维素和半纤维素作为唯一碳源的能力，并且能够直接发酵这些植物衍生的化合物以产生有价值的生物燃料，包括丁醇（Weselowski et al.，已提交和修订）。为了更好地理解多粘原杆菌CR 1代谢途径和涉及木质素降解和生物燃料生产的调控机制，我的实验室最近对其约6.02Mbp的完整基因组进行了测序和注释（Eastman et al.，2014. Genome Announcements）。我们最近的研究还表明，P. polymyxa CR 1可能利用了一种尚未确定的木质素和芳香族降解途径，因为它不能降解原儿茶酸，这是目前表征的细菌木质素降解途径的关键节点代谢产物。这与在所有测序的多粘原儿茶酸菌基因组中缺乏原儿茶酸裂解途径的鉴定同源物一致（Eastman等人，BMC Genomics. 2014）。* 在本提案中，基于我们最近产生的丰富的基因组和遗传信息以及我们在微生物遗传学方面的专业知识，我们将使用多种“组学”技术-包括功能基因组学，转录组学和代谢组学，以阐明P. polymyxa CR 1如何在系统和分子水平上识别和代谢木质素。 特别是，我们的目标是揭示直接涉及木质素降解和生物燃料生产的细菌基因、代谢途径和调控网络，这是一个先决条件，并将为我们的长期目标提供良好的基础，即对多粘原胞菌CR 1进行遗传工程改造，以提高木质素降解的效率，并改善直接从木质纤维素生物质生产生物燃料和增值化学品，从而使生物精炼具有成本效益并且生物燃料在经济上可行。 从拟议的研究中产生的知识，一旦被翻译，将在几个方面为生物燃料行业和生物经济做出重大贡献，包括降低生物炼制成本，增加生物燃料的数量和纯度，开发新的供应链和其他低价值木质纤维素原料的新市场，如林业/作物残留物，生物固体等。

Biofuels derived from lignocellulosic biomass hold promise as alternative sources of cleaner and renewable energy and chemicals. However, delignification or lignin decomposition is a bottleneck for biofuel and biorefinery. Therefore, it is very important to understand lignin degrading pathways, biofuel production and their regulatory mechanisms in bacteria. As part of my research program aiming at developing novel strategies for sustainable agriculture and environment, my lab recently characterized Paenibacilluspolymyxa CR1, a Gram-positive, sporulating, facultative anaerobe that we isolated from degrading corn roots in southern Ontario. We found P. polymyxa CR1 had a strong capacity to degrade and utilize lignin, cellulose and hemi-cellulose as a sole carbon source, and is able to ferment these plant-derived compounds directly to produce valuable biofuels including butanol (Weselowski et al., submitted and in revision). To better understand P. polymyxa CR1 metabolic pathways and regulatory mechanisms implicated in lignin degradation and biofuel production, my lab recently sequenced and annotated its complete genome which is approximately 6.02Mbp (Eastman et al., 2014. Genome Announcements). Our recent research also suggests P. polymyxa CR1 likely utilizes a yet to be identified lignin and aromatic degradation pathway, since it is incapable of degrading protocatechuate, a key nodal metabolite of currently characterized bacterial lignin degradation pathways. This is consistent with the lack of identified homologs for the protocatechuate cleavage pathway in all sequenced P. polymyxa genomes (Eastman et al., BMC Genomics. 2014). ***In this proposal, based on the wealth of genomic and genetic information we recently generated and our expertise of microbial genetics, we will use multiple "omics" techniques - including functional genomics, transcriptomics and metabolomics, to elucidate how P. polymyxa CR1 recognizes and metabolizes lignin at the system and molecular levels.  In particular, we aim to uncover bacterial genes, metabolic pathways and regulatory networks directly implicated in lignin degradation and biofuel production, which is a prerequisite and will provide a sound foundation for our long term goal of engineering P. polymyxa CR1 genetically to enhance the efficiency of lignin degradation and improve the production of biofuels and value-added chemicals directly from lignocellulosic biomass, thereby making biorefinery cost-efficient and biofuels economically viable.  Knowledge generated from the proposed study, once being translated, will significantly contribute to biofuel industry and bio-economy in several ways including reduced biorefinery cost, increased quantity and purity of biofuels, development of new supply chains and a new marketplace for otherwise low value lignocellulose feedstock such as forestry/crop residues, biosolids and etc. **