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KMID : 0545120160260040659
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Journal of Microbiology and Biotechnology
2016 Volume.26 No. 4 p.659 ~ p.665
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Identification of Oligosaccharides in Human Milk Bound onto the Toxin A Carbohydrate Binding Site of Clostridium difficile
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Nguyen Thi Thanh Hanh
Kim Jong-Woon
Park Jun-Seong
Hwang Kyeong-Hwan
Jang Tae-Su
Kim Chun-Hyung
Kim Do-Man
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Abstract
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The oligosaccharides in human milk constitute a major innate immunological mechanism by which breastfed infants gain protection against infectious diarrhea. Clostridium difficile is the most important cause of nosocomial diarrhea, and the C-terminus of toxin A with its carbohydrate binding site, TcdA-f2, demonstrates specific abolishment of cytotoxicity and receptor binding activity upon diethylpyrocarbonate modification of the histidine residues in TcdA. TcdA-f2 was cloned and expressed in E. coli BL21 (DE3). A human milk oligosaccharide (HMO) mixture displayed binding with TcdA-f2 at 38.2 respond units (RU) at the concentration of 20 μg/ml, whereas the eight purified HMOs showed binding with the carbohydrate binding site of TcdA-f2 at 3.3 to 14 RU depending on their structures via a surface plasma resonance biosensor. Among them, Lacto-N-fucopentaose V (LNFPV) and Lacto-N-neohexaose (LNnH) demonstrated tight binding to TcdA-f2 with docking energy of ?9.48 kcal/mol and ?12.81 kcal/mol, respectively. It displayed numerous hydrogen bonding and hydrophobic interactions with amino acid residues of TcdA-f2.
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KEYWORD
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Clostridium difficile, human milk oligosaccharides, molecular docking, surface plasmon resonance, toxin A
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