Abstract
BackgroundThe biodegradation of chitin is an important part of the carbon and nitrogen cycles in nature. Speeding up the biotransformation of chitin substrates can not only reduce pollution, but also produce high value-added products. However, this process is strictly regulated by the catalytic efficiency of the chitinolytic machinery. Therefore, it is necessary to study the mode of action and compound mechanisms of different chitin-degrading enzymes in depth to improve the catalytic efficiency of the chitinolytic machinery.ResultsThe thermophilic bacterium Streptomyces sp. F-3 showed comparatively high chitin degradation activities. To elucidate the mechanism underlying chitin hydrolysis, six chitin degradation-related enzymes were identified in the extracellular proteome of Streptomyces sp. F-3, including three chitinases (SsChi18A, SsChi18B, and SsChi18C) from the GH18 family, one GH19 chitinase (SsChi19A), one GH20 β-N-acetylhexosaminidase (SsGH20A), and one lytic polysaccharide monooxygenase (SsLPMO10A) from the AA10 family. All were upregulated by chitin. The heterologously expressed hydrolases could withstand temperatures up to 70 °C and were stable at pH values of 4 to 11. Biochemical analyses displayed that these chitin degradation-related enzymes had different functions and thus showed synergistic effects during chitin degradation. Furthermore, based on structural bioinformatics data, we speculated that the different action modes among the three GH18 chitinases may be caused by loop differences in their active site architectures. Among them, SsChi18A is probably processive and mainly acts on polysaccharides, while SsChi18B and SsChi18C are likely endo-non-processive and displayed higher activity on the degradation of chitin oligosaccharides. In addition, proteomic data and synergy experiments also indicated the importance of SsLPMO10A, which could promote the activities of the hydrolases and increase the monosaccharide content in the reaction system, respectively.ConclusionsIn this article, the chitinolytic machinery of a thermophilic Streptomyces species was studied to explore the structural basis for the synergistic actions of chitinases from different GH18 subfamilies. The elucidation of the degradation mechanisms of these thermophilic chitinases will lay a theoretical foundation for the efficient industrialized transformation of natural chitin.
Highlights
Chitin, a polymer of β-1,4-N-acetylglucosamine, is the second most abundant polysaccharide in nature after cellulose [1]
F-3 is able to withstand high temperatures, at which comparatively high activity of chitin degradation was observed, genes coding for chitin metabolism were analyzed
Three hydrolases belong to the GH18 family, they can be classified into three subfamilies based on their catalytic domains [19]: SsChi18A belonging to the sub-family A, SsChi18B from the sub-family B, and SsChi18C from the sub-family C (Fig. 1a)
Summary
A polymer of β-1,4-N-acetylglucosamine, is the second most abundant polysaccharide in nature after cellulose [1]. Chitin can be biotransformed into pharmacologically active products, such as N-acetylglucosamine and chitin oligosaccharides (CHOSs), which can be used as antimicrobial agents and immune enhancers to activate the host defense system These substances can be used as drug delivery vehicles and antioxidants, which could be useful in hemostasis, wound healing, blood cholesterol control, and food preservation [3], as well as have important anti-tumor and anti-infection activities [4]. Speeding up the biotransformation of chitin substrates can reduce pollution, and produce high value-added products. This process is strictly regulated by the catalytic efficiency of the chitinolytic machinery. It is necessary to study the mode of action and compound mechanisms of different chitin-degrading enzymes in depth to improve the catalytic efficiency of the chitinolytic machinery
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