Fig. 2
Budding of defined FFV five-alanine Gag mutants. a Five-alanine scanning mutagenesis at the N-terminus of FFV Gag. Eight Gag mutants (mLNPLQ, mLQQLY, mINNGL, mQPNPG, mHGDII, mAVRFT, mGGPWG and mPGDRW) were constructed based on the proviral clone pCF-7 or the Gag expression plasmid pBC-Gag-oPRE. m indicates a replacement of the original sequence with five alanines (underlined). b 293T cells were transfected in 10 cm dishes with 8 µg of each proviral pCF-7-based Gag mutant (mLNPLQ, mLQQLY, mINNGL, mQPNPG, mHGDII, mAVRFT, mGGPWG or mPGDRW) (lanes 1–8), pCF-7 (lane 9) or pcDNA (lane 10). Aliquots of cell lysates and VLPs in supernatants were analyzed by SDS-PAGE. Positions of Gag p48 and p52, full-length Env precursor gp130Env; mature processed Env gp48™, Pol precursor p127Pol and PR-RT-RN domain p65Pol are marked. c Two days p.t., released particles were titrated onto FeFab cells. Mean titers of three independent experiments are given. Error bars represent the standard deviation