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. 2008 Nov 19;83(3):1483–1491. doi: 10.1128/JVI.02057-08

FIG. 1.

FIG. 1.

The L240K ERp29 mutant does not stimulate PyV unfolding and infection efficiently. (A) DTT and EGTA destabilize PyV. PyV was incubated with DTT and EGTA where indicated, followed by the addition of trypsin. The samples were subjected to reducing SDS-PAGE followed by immunoblotting with an antibody against VP1. (B) L240K ERp29 cannot unfold VP1. PyV was preincubated with extracts from mock-transfected NIH 3T3 cells or from cells overexpressing WT, L240K, K226E, L227S, E220Q, or R223A ERp29. Trypsin was then added to the reaction mixtures. The samples were analyzed by reducing SDS-PAGE and immunoblotting with antibodies against VP1 (top panel) or ERp29 (bottom panel). The asterisk (*) represents a partial degradation product of VP1a. (C) L240K ERp29 stimulates PyV infection inefficiently. Mock-transfected NIH 3T3 cells and cells overexpressing WT or L240K ERp29 were challenged with PyV (100 PFU/cell). Large-T-antigen expression was analyzed by standard fluorescence microscopy. (D) The experiment was the same as for panel C, except that cells were challenged with 200 PFU/cell. Data are means ± standard deviations.