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Fig. 2 | BMC Medical Genetics

Fig. 2

From: Biochemical effects of mutations in the gene encoding the alpha subunit of eukaryotic initiation factor (eIF) 2B associated with Vanishing White Matter disease

Fig. 2

Biochemical effects of mutations of eIF2Bα on eIF2B complex formation. a HEK293 cells were transfected with plasmids encoding myc-tagged eIF2B subunits andHis6-myc-tagged eIF2Bα. ‘No His-tag’ indicates that a vector encoding myc-tagged eIF2Bα was used instead. Lysates from transfected cells were analysed by western blotting for the myc tag to verify even expression of each subunit. An actin loading control is included. b Lysates were subjected to affinity purification using Ni2+ agarose and the bound material was analysed by western blot for myc tag to test for the presence of other subunits associated with His6-mys-eIF2Bα and phospho- and total eIF2α to assess substrate binding. c The myc-signal for the WT and indicated mutations were quantified and normalised to the level of eIF2Bα. Data are shown relative to the level of the WT pulldown. Data are shown as the relative expression to WT ± SEM of 3 independent experiments. P-values are as indicated on the figure. d Levels of total and phospho-eIF2 from pulldowns were quantified and normalised to the level of eIF2Bα

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