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Fig. 4 | Virology Journal

Fig. 4

From: Study of SV40 large T antigen nucleotide specificity for DNA unwinding

Fig. 4

Oligomerization assay of LT WT and nucleotide pocket mutants using gel filtration chromatography on Superdex 200 column. af Gel filtration profiles of WT, cS430T, tK419A, cW393A, and cL557A without NTP (a) with ATP (b) with ATP + Mg++ (c) with TTP (d) with TTP + Mg++ (e) and with UTP ± Mg++ (f). The assays were performed by analyzing 250 μg of LT protein with or without the mentioned NTP and Mg++ using Superdex 200 10/300 GL column. The two peaks represent the hexamer (~344 kDa) and monomer (~57 kDa) of LT. g The percentages of hexamer formation which were calculated based on the areas under each UV absorption peak in panels A–F. “─” indicates that the oligomerization test in the presence of UTP ± Mg++ was not performed. The running buffer contained 25 mM Tris pH 8.0, 250 mM NaCl, and 0.5 mM TCEP. Each protein was incubated with 4 mM NTP and 1 mM MgCl2 where indicated

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