Characterization of V3 Loop-Binding Protein(s) of Molt-4 and U937 Cells

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Abstract
The V3 loop in gpl20 of human immunodeficiency virus type 1 (HIV-1) is known as a principal neutralizing and cell-tropic determinant. Biotinylated synthetic V3 loop peptides derived from three different HIV-1 strains were used as ligands to identify the cell surface counterreceptor, which may participate in the infection of HIV-1. Two different cell lines, Molt-4 and U937, and three V3 loop peptides derived from LAVELI, HTLVIIIMN, and HTLV-IIIB strains were used. The binding of HTLV-IIIB-derived peptide to the cell surface was confirmed using 125I-labeled surface proteins of both cell lines. The relative molecular mass of the major radioactive band on the autoradiogram was 32-33 kDa in both cell lines. A protein was purified from the plasma membrane fraction of Molt-4 cells using affinity columns coupled with three different V3 loop peptides. Two major polypeptides (32 and 33 kDa) were eluted from the affinity column. Size-exclusion chromatography showed that the protein migrated as a single peak with a molecular mass of 130 kDa. These proteins were separated by reversed-phase chromatography, which indicated that the 32-kDa protein is more hydrophobic than the 33-kDa protein in Molt-4 cells. A similar but not identical 130-kDa protein with 32- and 33-kDa polypeptides were also purified from U937 cells. These findings indicate that HIV-1 utilizes a tetrameric protein on the surface of Molt-4 and U937 cells on infection.