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Polymers 2018, 10(3), 237; https://doi.org/10.3390/polym10030237

Biosynthetic Pathway and Genes of Chitin/Chitosan-Like Bioflocculant in the Genus Citrobacter

1
Department of Applied Chemistry, Graduate School of Engineering, University of Hyogo, 2167 Shosha, Himeji, Hyogo 671-2280, Japan
2
Hyogo Analysis Center Co., Ltd., 4-10-8 Seimondori, Hirohata, Himeji, Hyogo 671-1116, Japan
3
Microbial Type Culture Collection & Gene Bank (MTCC), CSIR-Institute of Microbial Technology, Sector 39-A, Chandigarh-160 036, India
4
Department of Chemistry and Bioscience, Graduate School of Science and Engineering, Kagoshima 890-8580, Japan
*
Author to whom correspondence should be addressed.
Received: 23 January 2018 / Revised: 19 February 2018 / Accepted: 22 February 2018 / Published: 27 February 2018
(This article belongs to the Special Issue Advances in Chitin/Chitosan Characterization and Applications)
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Abstract

Chitin/chitosan, one of the most abundant polysaccharides in nature, is industrially produced as a powder or flake form from the exoskeletons of crustaceans such as crabs and shrimps. Intriguingly, many bacterial strains in the genus Citrobacter secrete a soluble chitin/chitosan-like polysaccharide into the culture medium during growth in acetate. Because this polysaccharide shows strong flocculation activity for suspended solids in water, it can be used as a bioflocculant (BF). The BF synthetic pathway of C. freundii IFO 13545 is expected from known bacterial metabolic pathways to be as follows: acetate is metabolized in the TCA cycle and the glyoxylate shunt via acetyl-CoA. Next, fructose 6-phosphate is generated from the intermediates of the TCA cycle through gluconeogenesis and enters into the hexosamine synthetic pathway to form UDP-N-acetylglucosamine, which is used as a direct precursor to extend the BF polysaccharide chain. We conducted the draft genome sequencing of IFO 13545 and identified all of the candidate genes corresponding to the enzymes in this pathway in the 5420-kb genome sequence. Disruption of the genes encoding acetyl-CoA synthetase and isocitrate lyase by homologous recombination resulted in little or no growth on acetate, indicating that the cell growth depends on acetate assimilation via the glyoxylate shunt. Disruption of the gene encoding glucosamine 6-phosphate synthase, a key enzyme for the hexosamine synthetic pathway, caused a significant decrease in flocculation activity, demonstrating that this pathway is primarily used for the BF biosynthesis. A gene cluster necessary for the polymerization and secretion of BF, named bfpABCD, was also identified for the first time. In addition, quantitative RT-PCR analysis of several key genes in the expected pathway was conducted to know their expression in acetate assimilation and BF biosynthesis. Based on the data obtained in this study, an overview of the BF synthetic pathway is discussed. View Full-Text
Keywords: Citrobacter; biosynthesis; bioflocculant; chitosan; metabolic pathway Citrobacter; biosynthesis; bioflocculant; chitosan; metabolic pathway
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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Takeo, M.; Kimura, K.; Mayilraj, S.; Inoue, T.; Tada, S.; Miyamoto, K.; Kashiwa, M.; Ikemoto, K.; Baranwal, P.; Kato, D.; Negoro, S. Biosynthetic Pathway and Genes of Chitin/Chitosan-Like Bioflocculant in the Genus Citrobacter. Polymers 2018, 10, 237.

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