Qu Wu, Lin Dan, Zhang Zhouhao, Di Wenjie, Gao Boliang, Zeng Runying
School of Life Sciences, Xiamen University, Xiamen, China.
Novogene Bioinformatics Technology Co. Ltd., Tianjin, China.
Front Microbiol. 2018 Aug 14;9:1864. doi: 10.3389/fmicb.2018.01864. eCollection 2018.
Monosaccharides and oligosaccharides produced by agarose degradation exhibit potential in the fields of bioenergy, medicine, and cosmetics. Mangrove sediments (MGSs) provide a special environment to enrich enzymes for agarose degradation. However, representative investigations of the agarlytic genes in MGSs have been rarely reported. In this study, agarlytic genes in MGSs were researched in detail from the aspects of diversity, abundance, activity, and location through deep metagenomics sequencing. Functional genes in MGSs were usually incomplete but were shown as results, which could cause virtually high number of results in previous studies because multiple fragmented sequences could originate from the same genes. In our work, only complete and nonredundant (CNR) genes were analyzed to avoid virtually high amount of the results. The number of CNR agarlytic genes in our datasets was significantly higher than that in the datasets of previous studies. Twenty-one recombinant agarases with agarose-degrading activity were detected using heterologous expression based on numerous complete open-reading frames, which are rarely obtained in metagenomics sequencing of samples with complex microbial communities, such as MGSs. Aga2, which had the highest crude enzyme activity among the 21 recombinant agarases, was further purified and subjected to enzymatic characterization. With its high agarose-degrading activity, resistance to temperature changes and chemical agents, Aga2 could be a suitable option for industrial production. The agarase ratio with signal peptides to that without signal peptides in our MGS datasets was lower than that of other reported agarases. Six draft genomes, namely, Clusters 1-6, were recovered from the datasets. The taxonomic annotation of these genomes revealed that Clusters 1, 3, 5, and 6 were annotated as sp., sp., Ignavibacteriales spp., and Polyangiaceae spp., respectively. Meanwhile, Clusters 2 and 4 were potential new species. All these genomes were first reported and found to have abilities of degrading various important polysaccharides. The metabolic pathway of agarose in Cluster 4 was also speculated. Our results showed the capacity and activity of agarases in the MGS microbiome, and MGSs exert potential as a repertory for mining not only agarlytic genes but also almost all genes of the carbohydrate-active enzyme family.
琼脂糖降解产生的单糖和寡糖在生物能源、医学和化妆品领域具有潜力。红树林沉积物（MGSs）提供了一个富集琼脂糖降解酶的特殊环境。然而，关于MGSs中琼脂糖降解基因的代表性研究鲜有报道。在本研究中，通过深度宏基因组测序，从多样性、丰度、活性和定位等方面对MGSs中的琼脂糖降解基因进行了详细研究。MGSs中的功能基因通常不完整，但作为结果显示出来，这可能导致以往研究中出现大量结果，因为多个片段化序列可能来自同一基因。在我们的工作中，只分析完整且无冗余（CNR）的基因，以避免出现大量结果。我们数据集中CNR琼脂糖降解基因的数量显著高于以往研究数据集。基于众多完整的开放阅读框，通过异源表达检测到21种具有琼脂糖降解活性的重组琼脂酶，而在具有复杂微生物群落的样本（如MGSs）的宏基因组测序中很少能获得这些开放阅读框。Aga2在21种重组琼脂酶中具有最高的粗酶活性，进一步对其进行纯化并进行酶学表征。由于其具有高琼脂糖降解活性、对温度变化和化学试剂的耐受性，Aga2可能是工业生产的合适选择。我们MGS数据集中有信号肽的琼脂酶与无信号肽的琼脂酶的比例低于其他已报道的琼脂酶。从数据集中获得了6个草图基因组，即聚类1 - 6。这些基因组的分类注释显示，聚类1、3、5和6分别被注释为 sp.、 sp.、Ignavibacteriales spp.和Polyangiaceae spp.。同时，聚类2和4是潜在的新物种。所有这些基因组均首次被报道，并发现具有降解各种重要多糖的能力。还推测了聚类4中琼脂糖的代谢途径。我们的结果显示了MGS微生物群落中琼脂酶的能力和活性，并且MGSs不仅作为挖掘琼脂糖降解基因的宝库，而且作为挖掘几乎所有碳水化合物活性酶家族基因的宝库具有潜力。
