Analogs of LDL Receptor Ligand Motifs in Dengue Envelope and Capsid Proteins as Potential Codes for Cell Entry

Figure 3

Effects of peptide E-2 and LDL on HeLa cell binding of Dsp2EP-FITC. (a) Unlabeled E-2 (40 µg, 160 µg, and 400 µg) was added to each culture containing 10 µg Dsp2EP in 500 µL PBS-5 mM MgCl₂ and placed at 37°C for 18 hours. Parallel assays were performed using unlabeled human LDL (300 µg, 600 µg, and 1200 µg) in lieu of E-2. Representative images obtained for experiments Dsp2EP-FITC alone, plus peptide E-2, and plus LDL are shown in frames (A), (B), and (C), respectively. Enlarged images (A1), (B1), and (C1) are of areas indicated in corresponding frames and show fluorescence signal in clusters on the cell surface. (b) Number of points (loci) of signal for images in frames (A), (B), and (C) of Figure 3(a). (c) total raw integrated densities for the same frames (A), (B), and (C) in Figure 3(a). Image J software was used to count loci and integrate the fluorescence signal. The number of fluorescing loci and integrated density values for cells treated with 10 µg Dsp2EP-FITC alone are shown in green columns, Dsp2EP-FITC plus unlabeled E-2 peptide is in blue, and Dsp2EP-FITC plus unlabeled purified human LDL are in orange. Amounts of E-2 peptide or LDL used in the assays are shown under each column. E-2 peptide at high concentration significantly enhances binding of Dsp2EP-FITC to HeLa cells which appears to be ubiquitous, frame (B) in Figure 3(a).

(a)

(b)

(c)