Abstract: In a previous study we noted significant THP binding to TNF-α, but did not explore the molecular basis of the structure-binding relationship. In this study, we used lectin-binding ELISA to assess the carbohydrate compositions of THP, BSA, IgG, TNF-α, and IFN-g. We identified β(1,4)-N-acetylglucosamine oligomers (GlcNAc) and GlcNAc/branched mannose in BSA, IgG, TNF-α, and THP, but not in IFN-g. These carbohydrate moieties mediated binding with THP. Small amounts of Siaα(2,3)Gal/ GalNAc, Sia(2,6)Gal/GalNAc, and mannose residues were also present in THP and TNF-α. Binding affinity (Kd) between THP and TNF-α by Scatchard plot analysis was 1.4–1.7 × 10−6 M, lower than antigen-antibody or ligand-receptor binding affinities. To elucidate the structure-binding relationship of THP-TNF-α, THP was digested with neuraminidase, β-galactosidase, O-sialoglycoprotein endopeptidase, carboxypeptidase Y, or proteinase K. β-galactosidase increased binding capacity of THP for TNF-α. Monosaccharide inhibition suggested that α-methyl-D-mannoside, GlcNAc, and GalNAc, but not sialic acid, suppress THP-TNF-α binding as detected by ELISA. We conclude that sugar-lectin and sugar-protein interactions between cognate sites in THP and TNF-α mediate their binding.
Keywords: Tamm-Horsfall glycoprotein; tumor necrosis factor-α; binding affinity; structure-binding relationship; glucosamine-containing mannose
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Wu, C.-H.; Li, K.-J.; Siao, S.-C.; Chen, Y.-H.; Wu, T.-H.; Tsai, C.-Y.; Yu, C.-L. The Binding Affinity and Molecular Basis of the Structure-Binding Relationship between Urinary Tamm-Horsfall Glycoprotein and Tumor Necrosis Factor-α. Molecules 2012, 17, 11978-11989.
Wu C-H, Li K-J, Siao S-C, Chen Y-H, Wu T-H, Tsai C-Y, Yu C-L. The Binding Affinity and Molecular Basis of the Structure-Binding Relationship between Urinary Tamm-Horsfall Glycoprotein and Tumor Necrosis Factor-α. Molecules. 2012; 17(10):11978-11989.
Wu, Cheng-Han; Li, Ko-Jen; Siao, Sue-Cien; Chen, Yu-Hsuan; Wu, Tsai-Hung; Tsai, Chang-Youh; Yu, Chia-Li. 2012. "The Binding Affinity and Molecular Basis of the Structure-Binding Relationship between Urinary Tamm-Horsfall Glycoprotein and Tumor Necrosis Factor-α." Molecules 17, no. 10: 11978-11989.