¹²⁵ J-ASA-PE, A Photoactivable, Radioactive Phospholipid-Analogue, Designed for Photolabeling Proteins in Contact with Phospholipid Head Groups

Abstract

To photolabel proteins, which are in close proximity to phospholipid head groups on membranes, N-(4-azidosalicyl)phosphatidylethanolamine, ASA-PE, was synthesized. Its special advantage is that it can be radioiodinated in the last step of the synthesis to the [¹²⁵J]-iodo compound (¹²⁵J-ASA-PE) with very high specific activity. This is necessary because the4-azido-3[¹²⁵J]iodo-2-hydroxybenzoyl group is located on the membrane surface and reacts during photolysis as well with proteins as with water. Additionally high specific activity allows to insert low amounts of ¹²⁵J-ASA-PE into proteoliposomes thus preserving biological activity of their proteins. To study structure and function of the thermophilic ATPsynthase TF_o·F₁ of the thermophilic bacterium PS 3, it was first reconstituted with soybean phospholipids into liposomes and these were then incubated with ¹²⁵J-ASA-PE to provide preferred incorporation into the outside of the phospholipid bilayer. After photolysis subunits α of TF_o·F₁ were labeled selectively. It is, therefore, possible that subunits a may be important in the interaction of TF₁ with TFo, to provide internal energy coupling between the proton channel TFo and ATP synthesis in TF₁.