Interaction of human β-defensin 2 (HBD2) with glycosaminoglycans.
Human β-defensin 2 (HBD2) is a member of the defensin family of antimicrobial peptides that plays important roles in the innate and adaptive immune system of both vertebrates and invertebrates. In addition to their direct bactericidal action, defensins are also involved in chemotaxis and Toll-like receptor activation. In analogy to chemokine/glycosaminoglycan (GAG) interactions, GAG-defensin complexes are likely to play an important role in chemotaxis and in presenting defensins to their receptors. Using a gel mobility shift assay, we found that HBD2 bound to a range of GAGs including heparin/heparan sulfate (HS), dermatan sulfate (DS), and chondroitin sulfate. We used NMR spectroscopy of (15)N-labeled HBD2 to map the binding sites for two GAG model compounds, a heparin/HS pentasaccharide (fondaparinux sodium; FX) and enzymatically prepared DS hexasaccharide (DSdp6). We identified a number of basic amino acids that form a common ligand binding site, which indicated that these interactions are predominantly electrostatic. The dissociation constant of the [DSdp6-HBD2] complex was determined by NMR spectroscopy to be 5 ± 5 μM. Binding of FX could not be quantified because of slow exchange on the NMR chemical shift time scale. FX was found to induce HBD2 dimerization as evidenced by the analysis of diffusion coefficients, (15)N relaxation, and nESI-MS measurements. The formation of FX-bridged HBD2 dimers exhibited features of a cooperative binding mechanism. In contrast, the complex with DSdp6 was found to be mostly monomeric.
EastChem, School of Chemistry, The University of Edinburgh, King's Buildings, West Mains Road, Edinburgh EH9 3JJ, UK.
Emily S
ES
Seo
Bärbel S
BS
Blaum
Thomas
T
Vargues
Martin
M
De Cecco
Jon A
JA
Deakin
Malcolm
M
Lyon
Perdita E
PE
Barran
Dominic J
DJ
Campopiano
Dusan
D
Uhrín
D001665:Binding Sites / Q000502:physiology; D002634:Chemotaxis, Leukocyte / Q000502:physiology; D006025:Glycosaminoglycans / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D055672:Static Electricity; D013431:Sulfates / Q000737:chemistry / Q000378:metabolism; D023083:beta-Defensins / Q000737:chemistry* / Q000378:metabolism*
C106641:DEFB4A protein, human; D006025:Glycosaminoglycans; D009844:Oligosaccharides; D013431:Sulfates; D023083:beta-Defensins