Interleukin-8 binds to syndecan-2 on human endothelial cells.
Application of reverse transcription-PCR to total RNA prepared from TNF-alpha (tumour necrosis factor-alpha)-stimulated HUVECs (human umbilical vein endothelial cells) revealed that the syndecan-2 mRNA was up-regulated by this inflammatory stimulus. By immunoprecipitation using an anti-syndecan-2 antibody on TNF-alpha-stimulated HUVEC lysates, inflammation-induced interleukin-8 was found to be an interaction partner of this HS (heparan sulphate) proteoglycan, but not of any other syndecan on these cells. The glycosylated [Syn2(ect)(+HS)] and non-glycosylated [Syn2(ect)(-HS)] forms of Syn2(ect) (the syndecan-2 ectodomain) were purified from a stably transfected human cell line and from a bacterial expression system respectively. By CD spectroscopy, Syn2(ect) was found to adopt an all-beta secondary structure. The dissociation constant of Syn2(ect)(+HS) with respect to interleukin-8 binding was determined by isothermal fluorescence titrations to be 23 nM. Despite its lack of HS chains, Syn2(ect)(-HS) exhibited significant binding to the chemokine, with a K (d) of >1 microM. Thus, in addition to glycosaminoglycan binding, protein-protein contacts might also contribute to the chemokine-proteoglycan interaction.
Halden
Yvonne
Y
Institute of Pharmaceutical Chemistry and Pharmaceutical Technology, University of Graz, Universitätsplatz 1, A-8010 Graz, Austria.
Rek
Angelika
A
Atzenhofer
Werner
W
Szilak
Laszlo
L
Wabnig
Astrid
A
Kungl
Andreas J
AJ
D001665:Binding Sites; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D017433:Protein Structure, Secondary; D011509:Proteoglycans / Q000737:chemistry / Q000378:metabolism*; D053669:Syndecan-2; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D016209:Interleukin-8; D008562:Membrane Glycoproteins; D011509:Proteoglycans; C508245:SDC2 protein, human; D014409:Tumor Necrosis Factor-alpha; D053669:Syndecan-2