Definition of a major p53 binding site on Ad2E1B58K protein and a possiblenuclear localization signal on the Ad12E1B54K protein

Previous studies have established that adenovirus 2/5 early region 1B (Ad E 1B) 58K protein binds p53 strongly and co-localizes with it to cytoplasmic dense bodies whilst the homologous Ad12E1B54K protein binds only weakly and co-localizes primarily to the nucleus in Ad12E1 transformed cells. We have used these properties of the E1B proteins from different viral serotypes t o map the p53 binding site on the Ad2/5 protein. A set of chimaeric genes w as constructed containing different proportions of the Ad12 and Ad2E1B DNA, These, together with Ad12E1A and E1B19K DNA, were transfected into baby ra t kidney cells and transformed lines isolated, From an examination of the p roperties of these Ad12/Ad2E1B fusion proteins in co-immunoprecipitation an d subcellular localization experiments it has been concluded that the p53 b inding site on Ad2E1B58K protein lies between amino acids 216 and 235 and t hat the homologous region on Ad12E1B54K protein also binds p53. In addition , a unique nuclear localization signal is located on Ad12E1B54K between res idues 228 and 239, We suggest that primary structure differences in these r egions of the Ad2 and Ad12E1B proteins are responsible for the different su bcellular localizations in AdE1 transformants.