VLDL receptor fragments of different lengths bind to human rhinovirus HRV2 with different stoichiometry. An analysis of virus-receptor complexes by capillary electrophoresis.
The formation of complexes between the minor receptor group human rhinovirus HRV2 and two recombinant soluble receptor fragments derived from the human very low density lipoprotein receptor (VLDLR) and containing ligand-binding repeats 1-3 (MBP.VLDLR(1-3)) or 1-8 (MBP.VLDLR(1-8)) fused to the carboxyl terminus of the maltose-binding protein was analyzed by affinity capillary electrophoresis. At low molar ratios of receptor/virus, the peaks corresponding to substoichiometric complexes were broad indicating heterogeneity. When the receptors were present in molar excess with respect to the virus, the peaks were sharp, suggesting saturation of all binding sites. For the determination of the stoichiometry, constant amounts of receptor were incubated with increasing amounts of virus, and the peak areas corresponding to free receptor were measured and plotted versus total virus concentration. Extrapolation of the linear part of the resulting curve to zero concentration of free receptor enabled quantitation of the molar ratios of the components present in the complex. Using this method, we determined that about 60 molecules of MBP.VLDLR(1-3) but only about 30 molecules of MBP.VLDLR(1-8) were bound per virion.
Okun
V M
VM
Institute of Medical Biochemistry, Vienna Biocenter, University of Vienna, Dr. Bohr Gasse 9/3, A-1030 Vienna, Austria.
Moser
R
R
Ronacher
B
B
Kenndler
E
E
Blaas
D
D
D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism; D019075:Electrophoresis, Capillary / Q000379:methods; D006801:Humans; D007700:Kinetics; D008079:Lipoproteins, VLDL / Q000378:metabolism; D058619:Maltose-Binding Proteins; D011973:Receptors, LDL / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012229:Rhinovirus / Q000145:classification / Q000235:genetics / Q000378:metabolism*
D002352:Carrier Proteins; D008079:Lipoproteins, VLDL; D058619:Maltose-Binding Proteins; D011973:Receptors, LDL; D011993:Recombinant Fusion Proteins; C036053:VLDL receptor