Hydrolysis of insoluble collagen by deseasin MCP-01 from deep-sea Pseudoalteromonas sp. SM9913: collagenolytic characters, collagen-binding ability of C-terminal polycystic kidney disease domain, and implication for its novel role in deep-sea sedimentary particulate organic nitrogen degradation |
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Authors: | Zhao Guo-Yan Chen Xiu-Lan Zhao Hui-Lin Xie Bin-Bin Zhou Bai-Cheng Zhang Yu-Zhong |
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Institution: | State Key Lab of Microbial Technology, Marine Biotechnology Research Center, Shandong University, Jinan 250100, China |
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Abstract: | Collagens are the most abundant proteins in marine animals and their degradation is important for the recycling of marine nitrogen. However, it is rather unclear how marine collagens are degraded because few marine collagenolytic proteases are studied in detail. Deseasins are a new type of multidomain subtilases. Here, the collagenolytic activity of deseasin MCP-01, the type example of deseasins, was studied. MCP-01 had broad substrate specificity to various type collagens from terrestrial and marine animals. It completely decomposed insoluble collagen into soluble peptides and amino acids, and was more prone to degrade marine collagen than terrestrial collagen. Thirty-seven cleavage sites of MCP-01 on bovine collagen chains were elucidated, showing the cleavage is various but specific. As the main extracellular cold-adapted protease from deep-sea bacterium Pseudoalteromonas sp. SM9913, MCP-01 displayed high activity at low temperature and alkaline range. Our data also showed that the C-terminal polycystic kidney disease (PKD) domain of MCP-01 was able to bind insoluble collagen and facilitate the insoluble collagen digestion by MCP-01. Site-directed mutagenesis demonstrated that Trp-36 of the PKD domain played a key role in its binding to insoluble collagen. It is the first time that the structure and function of a marine collagenolytic protease, deseasin MCP-01, has been studied in detail. Moreover, the PKD domain was experimentally proven to bind to insoluble protein for the first time. These results imply that MCP-01 would play an important role in the degradation of deep-sea sedimentary particulate organic nitrogen. |
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