VLDL receptor fragments of different lengths bind to human rhinovirus HRV2with different stoichiometry - An analysis of virus-receptor complexes by capillary electrophoresis
Vm. Okun et al., VLDL receptor fragments of different lengths bind to human rhinovirus HRV2with different stoichiometry - An analysis of virus-receptor complexes by capillary electrophoresis, J BIOL CHEM, 276(2), 2001, pp. 1057-1062
The formation of complexes between the minor receptor group human rhinoviru
s HRV2 and two recombinant soluble receptor fragments derived from the huma
n very low density lipoprotein receptor (VLDLR) and containing ligand-bindi
ng repeats 1-3 (MBP.VLDLR1-3) or 1-8 (MBP.VLDLR1-8) fused to the carboxyl t
erminus of the maltose binding protein was analyzed by affinity capillary e
lectrophoresis. At low molar ratios of receptor/virus, the peaks correspond
ing to substoichiometric complexes were broad indicating heterogeneity. Whe
n the receptors were present in molar excess with respect to the virus, the
peaks were sharp, suggesting saturation of all binding sites. For the dete
rmination 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. Extra
polation of the linear part of the resulting curve to zero concentration of
free receptor enabled quantitation of the molar ratios of the components p
resent in the complex. Using this method, we determined that about 60 molec
ules of MBP.VLDLR1-3 but only about 30 molecules of MBP.VLDLR1-8 were bound
per virion.