Fluidity Parameters of Lipid Regions Determined by Fluorescence Polarization

Mechanism of the Na+, K+ pump protein structure and conformations of the pure (Na+ + K+)-ATPase

Publisher Summary Studies of receptor binding generally employ highly radioactive ligand derivatives; however, knowledge of the biological activity of the radioactive derivative is essential for a complete interpretation of the binding data. This chapter describes the relation between biological activity and binding, physicochemical detection and assay of receptors, solubilization of membrane receptors, provides examples of membrane-localized receptors, affinity techniques, polyvalent hormone derivatives, and receptor studies, and theoretical mechanisms of hormone–receptor binding and action. The biological response toward many neurotransmitters and hormones, such as insulin, acetylcholine, and epinephrine is rapid both in onset and in the termination of action on removal of these agents; therefore, it may be anticipated that the initial hormone–receptor interaction will also be reversible. The chapter shows that progress has been made toward understanding the way by which hormones, via interactions with cell–surface receptors, modulate overall cellular function. This interaction is only the first in a presumably complicated series of protein–protein interactions within the plane of the cell membrane; the mechanism, whereby the ultimate signal is transmitted to the internal cellular milieu is undoubtedly an equally intricate process. The chapter is considered to be useful in delineating the challenge that the elucidation of these processes presents and in providing a basis and incentive for more discoveries in this field.

Membrane receptors and hormone action.

Publisher Summary This chapter discusses the photoaffinity labeling which could be used as a method that allows to unleash the reagent at a particular time and place, when the chemical affinity labeling restricts it. The chapter notes that the possibility that a labile group of appropriate reactivity cannot be incorporated into the ligand molecule without excessive disturbance of the recognition process, there are two limitations to the affinity labeling approach. The first challenge; the range of chemical reactivity of groups that can be incorporated into the ligand is limited by the fact that these groups must not react so rapidly with water that they are destroyed hydrolytically before the ligand that carries them can reach the binding site. And secondly, it is becoming clear that some biological problems require a reagent whose reactivity remains masked until the experimenter chooses to activate it. Both of the two limitations of classical chemical affinity labeling discussed above can in principle be circumvented by the use of a photogenerated reagent.

[8] Photoaffinity labeling

The synthesis of a new photoactivatable probe, 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine ([125I]-TID), with a high specific radioactivity (10 Ci mmol-1) is described. It was tested as a probe for the hydrophobic core of membranes. TID partitions strongly in favor of the lipid phase of membranes, and the photogenerated carbene labels intrinsic membrane proteins in a highly selective manner. This conclusion was reached from the distribution of radioactivity among the proteins of [125I]TID-labeled human erythrocyte membranes. By far the most heavily labeled protein is band 3 [nomenclature of Fairbanks, G., Steck, T. L., & Wallach, F. G. H. (1971) Biochemistry 10, 2606-2617] while the labeling of glycophorin is approximately 5 times less than that of band 3. There is little or no labeling of known extrinsic proteins.

Amira Klip

Papers

Photoactive covalent labeling of membrane components from within the lipid core.

Labelling of rhodopsin moieties confined to the membrane lipid bilayer