We have reported previously that glycoglycerolipids derived from the membranes of Acholeplasma laidlawii, 3-O-[2'-O-(alpha-D-glucopyranosyl)-6'-O-acyl-alpha-glucopyranosy]1-1,2-d i-O-acyl-sn-glycerols (GAGDGs) bind to human cell lines. In addition, the GAGDGs were found to augment HIV-1 infection in human cell lines. Here we show that GAGDG binds to HIV-1 and facilitates the entry of HIV-1 into cells. The binding ability of GAGDG to HIV-1 was blocked by anti-GAGDG serum. Binding assay with synthetic GAGDGs and related compounds showed that the presence of branching form of acyl chains at the C14 or C16 position, glucose, and the acyl chain binding to the glucose were critical for efficient binding. GAGDG efficiently augmented the entry of HIV-1 into cells in a single-cycle replication assay. These results indicate that GAGDG of A. laidlawii membranes participates in the facilitation of HIV-1 infection.