Background: DC-SIGN and related DC-SIGNR bind high mannose oligosaccharides on HIV-1 viral particle envelope glycoprotein gp120 enhancing virus infection efficiency by a mechanism that is not completely understood. DC-SIGN is a C-type lectin containing a short intracellular signaling domain, a transmembrane domain, a repeat domain, which contains 7.5 copies of a 23 amino acid repeat, and a functional carbohydrate recognition domain (CRD). Previously, we have shown that by regulating the number of repeat domains included with the CRD we are able to modulate receptor oligomerization from monomer, to dimer and octomer. We have also shown that fully oligomerized DC-SIGNR binds more avidly to HIV-1 gp120 IIIB than monomer or dimer species as seen by an increase in dissociation constant (K_D) from 10–20 nM for oligomer to ~10 mM for monomer species. We continue our study of DC-SIGNR recognition and binding by investigating gp120 glycosylation dependency and DC-SIGNR oligomers binding to ICAMs.

Methods: DC-SIGNR protein was expressed and purified using a recombinant bacterial system. Surface plasmon resonance binding measurements were obtained using a Biacore 3000 instrument. Deglycosylation of gp120 was performed by treatment with N-Glycosidae F.

Results: In contrast to previous binding results with gp120 and DC-SIGNR oligomers, we observe that the DC-SIGNR binding to ICAM-3 is, however, not affected by receptor oligomerization and retains a relatively low binding affinity (11-50mM K_D) regardless of receptor oligomerization state. We also show that upon deglycosylation of gp120 binding to DC-SIGNR oligomer is abolished, while denatured fully glycosylated gp120 retains similar high affinity binding to DC-SIGNR oligomer as observed with native gp120. In addition to biochemical studies aimed at defining the DC-SIGNR and gp120 interaction, we have solved the 3-dimensional structure of DC-SIGNR’s carbohydrate binding domain with a one-half repeat.

Conclusions: The denaturation and deglycosylation gp120 binding results indicate that like ICAM-3 binding DC-SIGNR recognizes primarily the glycosylation state of gp120 and not additional structural features. Furthermore, the low affinity interaction of DC-SIGNR oligomers with ICAM-3 in contrast to the high affinity interaction with gp120 indicate DC-SIGN’s role is primarily antigen binding. Understanding the mechanism of DC-SIGNR’s binding of viral envelope glycoproteins, like gp120, may lead to a general understanding of viral particle host integration, as well provide new vaccine and drug therapies aimed at preventing viral infection.