FIG. 2.
Effects of proline residues on the protein stability of Vpr. (A) In transfected HeLa cells, protein biosynthesis was blocked. Samples were taken at the times indicated, and cell lysates were subjected to Western blotting. (B) Transfected cells were pulse-labeled with [35S]methionine for 10 min and chased for the times indicated. After cell lysis, FLAG-Vpr was immunoprecipitated with anti-FLAG antibodies, separated by 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and analyzed by fluorography. (C) The Vpr proteins for which results are shown in panel B were quantified with an image analyzer. The percentages of wt Vpr and proline mutants recovered relative to the amounts present at the end of the pulse (0 h) were plotted as a function of time.