出版社：American Society for Biochemistry and Molecular Biology
摘要：In order to have a model compound for detection of proteins involved in transport and metabolism of long-chain fatty acid salts by photoaffinity labeling 11,11-azistearate and 11,11-azi[G-3H]stearate (specific radioactivity 2.78 TBq/mmol) were synthesized. The suitability of 11,11-azi[G-3H]stearate for photoaffinity labeling was demonstrated by incorporation into BSA (bovine serum albumin) and H-FABP (hepatic fatty acid salt-binding protein) of rat liver. Repeated photoaffinity labeling resulted in a clear decrease of the binding capacities of both proteins. Labeling of protein mixtures with 11,11-azi[G-3H]stearate showed that binding proteins for long-chain fatty acid salts interact specifically with this probe. Photoaffinity labeling of isolated hepatocytes using 300 microM 11,11-azistearate in the presence of 100 microM BSA resulted in the irreversible inhibition of the uptake of stearate and its analogue 2,2,3,3,18,18,18-heptafluorostearate nearly to the same extent of about 30%. Irreversible inhibition of the uptake of long-chain fatty acid salts by photoaffinity labeling did not alter the mediated transport of cholyltaurine and has no effect on the uptake of 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol, a compound that crosses the hepatocyte membrane by simple diffusion. The irreversible inhibition of membrane transport by photoaffinity labeling demonstrates the existence of a specific transport system for the uptake of long-chain fatty acid salts into hepatocytes.
