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Title (Primary) Investigation of lysine side chain interactions of interleukin-8 with heparin and other glycosaminoglycans studied by a methylation-NMR approach
Author Möbius, K.; Nordsieck, K.; Pichert, A.; Samsonov, S.A.; Thomas, L.; Schiller, J.; Kalkhof, S.; Pisabarro, M.T.; Beck-Sickinger, A.G.; Huster, D.;
Journal Glycobiology
Year 2013
Department PROTEOM;
Volume 23
Issue 11
Language englisch;
POF III (all) F11;
Keywords chemokine;electrostatics;NMR;protein/glycosaminoglycan interaction;reductive methylation
UFZ wide themes ru3
Abstract

Although the interaction between interleukin-8 (IL-8) and glycosaminoglycans (GAGs) is crucial for the mediation of inflammatory effects, little is known about the site specificity of this interaction. Therefore, we studied complexes of IL-8 and heparin (HEP) as well as other GAGs in a multidisciplinary approach, involving site-directed mutagenesis, mass spectrometry, fluorescence and solution NMR spectroscopy as well as computer modeling. The interaction between GAG and IL-8 is largely driven by the amine groups of the lysine and the guanidinium groups of arginine side chains. However, due to fast exchange with the solvent, it is typically not possible to detect NMR signals of those groups. Here, we applied reductive 13C-methylation of the lysine side chains providing sensitive NMR probes for monitoring directly the sites of GAG interaction in 1H-13C correlation experiments. We focused on the lysine side chains K25, K28, K59, K69 and K72 of IL-8 (1–77), which were reported to be involved in the binding to GAGs. The NMR signals of these residues were assigned in 1H-13C HSQC spectra through the help of site-directed mutagenesis. NMR and fluorescence titration experiments in combination with molecular docking and molecular dynamics simulations were applied to investigate the involvement of each lysine in the binding with HEP and various GAG hexasaccharides. We identified K25, K69 and K72 to be the most relevant binding anchors of IL-8(1–77) for the analyzed GAGs.

ID 14059
Persistent UFZ Identifier https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=14059
Möbius, K., Nordsieck, K., Pichert, A., Samsonov, S.A., Thomas, L., Schiller, J., Kalkhof, S., Pisabarro, M.T., Beck-Sickinger, A.G., Huster, D. (2013):
Investigation of lysine side chain interactions of interleukin-8 with heparin and other glycosaminoglycans studied by a methylation-NMR approach
Glycobiology 23 (11), 1260 - 1269