Chitin is an abundant waste product from shrimp and mushroom industries and as such, an appropriate secondary feedstock for biotechnological processes. However, chitin is a crystalline substrate embedded in complex biological matrices, and, therefore, difficult to utilize, requiring an equally complex chitinolytic machinery. Following a bottom-up approach, we here describe the step-wise development of a mutualistic, non-competitive consortium in which a lysine-auxotrophic Escherichia coli substrate converter cleaves the chitin monomer N-acetylglucosamine (GlcNAc) into glucosamine (GlcN) and acetate, but uses only acetate while leaving GlcN for growth of the lysine-secreting Corynebacterium glutamicum producer strain. We first engineered the substrate converter strain for growth on acetate but not GlcN, and the producer strain for growth on GlcN but not acetate. Growth of the two strains in co-culture in the presence of a mixture of GlcN and acetate was stabilized through lysine cross-feeding. Addition of recombinant chitinase to cleave chitin into GlcNAc₂, chitin deacetylase to convert GlcNAc₂ into GlcN₂ and acetate, and glucosaminidase to cleave GlcN₂ into GlcN supported growth of the two strains in co-culture in the presence of colloidal chitin as sole carbon source. Substrate converter strains secreting a chitinase or a β-1,4-glucosaminidase degraded chitin to GlcNAc₂ or GlcN₂ to GlcN, respectively, but required glucose for growth. In contrast, by cleaving GlcNAc into GlcN and acetate, a chitin deacetylase-expressing substrate converter enabled growth of the producer strain in co-culture with GlcNAc as sole carbon source, providing proof-of-principle for a fully integrated co-culture for the biotechnological utilization of chitin.

Graphical abstract

Key Points

• A bacterial consortium was developed to use chitin as feedstock for the bioeconomy.

• Substrate converter and producer strain use different chitin hydrolysis products.

• Substrate converter and producer strain are mutually dependent on each other.

中文翻译：

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自下而上的方法对细菌财团进行几丁质向l-赖氨酸的生物技术转化

甲壳质是虾和蘑菇产业的一种丰富的废物，因此，它是生物技术过程的合适辅助原料。然而，甲壳质是嵌入复杂生物基质中的结晶底物，因此难以利用，需要同样复杂的壳多糖分解机制。按照自下而上的方法，在此我们描述了一个相互竞争的，非竞争性的财团的逐步发展，其中赖氨酸-营养缺陷型大肠杆菌底物转化体将几丁质单体N-乙酰基葡萄糖胺（GlcNAc）裂解为葡萄糖胺（GlcN）和乙酸盐，但仅使用乙酸盐，而将GlcN留在分泌赖氨酸的谷氨酸棒杆菌的生长中生产者菌株。我们首先设计了底物转化菌株，使其在乙酸盐上生长而不是在GlcN上生长，而生产菌株则在GlcN上生长但不在乙酸盐上生长。在GlcN和乙酸盐的混合物存在下，两种菌株在共培养中的生长通过赖氨酸交叉进料而稳定。加入重组几丁质酶将几丁质裂解为GlcNAc ₂，几丁质脱乙酰基酶将GlcNAc ₂转化为GlcN ₂和乙酸酯，以及氨基葡萄糖苷酶将GlcN ₂裂解为GlcN，这支持了两种菌株在胶体几丁质作为唯一碳存在下的共培养资源。分泌几丁质酶或β-1,4-氨基葡萄糖苷酶降解的几丁质的底物转化菌株为GlcNAc ₂或GlcN ₂分别为GlcN，但需要葡萄糖才能生长。相比之下，通过将GlcNAc裂解为GlcN和乙酸盐，表达几丁质脱乙酰基酶的底物转化子可以使生产菌株在与GlcNAc作为唯一碳源的共培养中生长，从而为完全整合的共培养提供了原理证明。甲壳素的生物技术利用。

图形概要

关键点

•建立了一个细菌联盟，以几丁质作为生物经济的原料。

•底物转化者和生产者菌株使用不同的几丁质水解产物。

•基板转换器和生产者应变相互依赖。