Lipid-protein interactions in monolayers

Abstract

We restricted ourselves to a few examples of the different methodological aspects of the investigation of lipid-protein interactions in monolayer assemblies. Experiments with monolayers have the unique advantage that the arrangement and packing of the molecules can be easily measured and controlled. The first section is devoted to proteins which do not degrade lipids. Soluble proteins are generally injected in the subphase below a preformed lipid monolayer and measurements are performed either at constant surface area or at constant surface pressure. These experiments can give information on the penetration capacity into the interface, its lipid specificity with a direct access to the area of the protein segment interacting with lipids. Reconstitution of functional enzymatic complexes can be achieved as well as the determination of the orientation of the protein at the interface. Most intrinsic membrane proteins are insoluble in water. In the absence of detergent they aggregate and display no affinity for lipid interfaces. These proteins can be spread from an organic solvent solution but with the risk of being denatured. In order to circumvent this difficulty a method for spreading an aqueous suspension of lipoproteins or natural membrane vesicles was developed. This spreading method allows the formation of lipoprotein films retaining biological activities and native membrane constituents. Formation of functional transmembrane complexes in planar bilayers from such lipoprotein films is the most fascinating application of this spreading technique. In the second section, we reviewed the use of pure or mixed lipid monolayers as substrates for lipolytic enzymes. Either long-chain lipids were used and their surface density was not controlled; or short-chain lipids were applied, again without control of surface density; or short-chain lipids were used at constant surface density. Recently a method was developed to study the hydrolysis of long chain lipids with control of surface density. The monolayer technique allows us to study accurately the influence of surface pressure and protein cofactors on the hydrolysis velocity and lag time in lipolysis. Two types of mixed lipid monolayers can be formed at the air/water interface: either by spreading, from a volatile organic solvent, a mixture of water insoluble lipids or by injecting a detergent solution into the water subphase covered with a preformed pure lipid monolayer. These techniques are ideally suited for the study of the mode of action of lipolytic enzymes on controlled mixed interfaces.