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. Author manuscript; available in PMC: 2009 Mar 20.
Published in final edited form as: Biochemistry. 2008 Sep 5;47(39):10314–10323. doi: 10.1021/bi801006s

Figure 2.

Figure 2

EGF-dependent dimerization and kinase activation of purified, detergent-soluble EGFR. (a) Δ998-EGFR was expressed by transient transfection in 293S GntI−/− cells and purified from detergent-solubilized cell lysates by streptactin–Sepharose affinity chromatography in the presence of 0.1% Triton X-100. Lanes 1−5 show 5 μL of the corresponding 0.5 mL fractions eluted with 2.5 mM desthiobiotin and subjected to reducing SDS–PAGE and Coomassie blue staining. (b, c) Gel filtration chromatograms of EGFR samples from lane 3 in (a) treated (b) without or (c) with 20 μM EGF and subjected to gel filtration on Superose 6HR equilibrated in 0.4 mM dodecyl maltoside. Protein elution was monitored by absorbance at 280 nm (solid line), and kinase activity of eluate fractions was determined by monitoring the incorporation of 32P into an exogenous peptide substrate (dashed line, circles).