Colorectal Mucus Binds DC-SIGN and Inhibits HIV-1 Trans-Infection of CD4+ T-Lymphocytes
Fig 4
Biochemical analysis of the DC-SIGN binding component in CM.
(A) Unfractionated CM (input) or <30, 30–100 or >100kDa CM fractions were coated to an ELISA plate and DC-SIGN-Fc binding was determined. Binding of unfractionated CM is set to 100% and binding of the fractions is expressed as a relative percentage. The <30kDa fraction shows no DC-SIGN binding, between 30–100kDa shows limited binding whilst >100kDa shows stronger binding. (B) DC-SIGN-Fc was incubated with untreated, heated (10 min at 95°C), proteinase K treated CM or medium (negative control) prior to addition to a gp140 coated plate. Compared to the media alone control incubating DC-SIGN-Fc with treated or untreated CM led to similar reductions in gp140 binding. (C) DC-SIGN-Fc was incubated with CM, BSA or mannan prior to addition to a gp140 coated plate. Untreated, both CM and mannan inhibit DC-SIGN-Fc from binding gp140 compared to BSA (negative control). Depletion of mannose structures from CM, BSA and mannan by a pull-down with Galanthus Nivalis lectin does not alter the DC-SIGN-Fc binding capacity of CM while mannan loses its ability to prevent gp140 binding by DC-SIGN-Fc. Data points were performed in triplicate.