From the Department of Biological Sciences, University of Maryland, Baltimore County, Baltimore, Maryland 21250 and.
the Faculty of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, 1430 Ås, Norway.
J Biol Chem. 2018 Mar 9;293(10):3849-3859. doi: 10.1074/jbc.RA117.000849. Epub 2018 Jan 24.
Understanding the strategies used by bacteria to degrade polysaccharides constitutes an invaluable tool for biotechnological applications. Bacteria are major mediators of polysaccharide degradation in nature; however, the complex mechanisms used to detect, degrade, and consume these substrates are not well-understood, especially for recalcitrant polysaccharides such as chitin. It has been previously shown that the model bacterial saprophyte is able to catabolize chitin, but little is known about the enzymatic machinery underlying this capability. Previous analyses of the genome and proteome indicated the presence of four glycoside hydrolase family 18 (GH18) enzymes, and studies of the proteome indicated that all are involved in chitin utilization. Using a combination of and approaches, we have studied the roles of these four chitinases in chitin bioconversion. Genetic analyses showed that only the gene product is essential for the degradation of chitin substrates. Biochemical characterization of the four enzymes showed functional differences and synergistic effects during chitin degradation, indicating non-redundant roles in the cell. Transcriptomic studies revealed complex regulation of the chitin degradation machinery of and confirmed the importance of Chi18D and LPMO10A, a previously characterized chitin-active enzyme. With this systems biology approach, we deciphered the physiological relevance of the glycoside hydrolase family 18 enzymes for chitin degradation in , and the combination of and approaches provided a comprehensive understanding of the initial stages of chitin degradation by this bacterium.
了解细菌降解多糖的策略是生物技术应用的宝贵工具。细菌是自然界中多糖降解的主要介导者；然而，用于检测、降解和消耗这些底物的复杂机制还不太清楚，特别是对于甲壳素等难降解的多糖。先前的研究表明，模式细菌腐生菌 能够分解甲壳素，但对于这种能力的酶机制知之甚少。先前对 基因组和蛋白质组的分析表明存在四种糖苷水解酶家族 18（GH18）酶，而对蛋白质组的研究表明所有这些酶都参与了甲壳素的利用。我们使用 和 方法的组合，研究了这四种几丁质酶在几丁质生物转化中的作用。遗传分析表明，只有 基因产物对于降解几丁质底物是必需的。对四种酶的生化特性分析表明，它们在几丁质降解过程中具有功能差异和协同作用，表明在细胞中具有非冗余的作用。转录组学研究揭示了 几丁质降解机制的复杂调控，并证实了 Chi18D 和 LPMO10A 的重要性，LPMO10A 是一种以前被表征的几丁质活性酶。通过这种系统生物学方法，我们揭示了糖苷水解酶家族 18 酶在 中降解几丁质的生理相关性，并结合 和 方法全面了解了该细菌降解几丁质的初始阶段。
