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In vitro protein acyltransferase (PAT) assay Palmitoyl acyltransferase constituted by purified recombinant DHHC9 and GCP16 proteins (100 ng for each) were incubated with GLUT1 (2 g) in 25 l of reaction buffer (50 mM Tris-HCl [pH 7.4], 10 M palmitoyl alkyne-CoA, 1 M palmostatin B) at 25 C for 1 h followed by a Cu(I)-assisted click reaction with biotin picolyl azide (50 M) to biotinylate the proteins with the incorporation of palmitoyl alkyne

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The cytosolic NADP redox state is typically considered to be highly reduced, with NADPH/NADP + ratios of 50100:1 being standard textbook values 54,55

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