Membrane proteins are endowed with a wide variety of vital biological functions. The main aim of this research is to study their general structures in membrane and, in particular, interactions with phospholipids. Determination of amino acid sequences as far as possible or necessary will be carried out. The main approach for studying the tertiary structures within and outside the membranes will involve the use of phospholipids containing photosensitive groups. The latter, on photolysis, result in cross-linking to adjacent groups. Previously initiated studies on fatty acids containing photoactivable groups and their incorporation into phospholipids will be continued and extended. In addition, phospholipids containing photosensitive groups in polar head groups will be synthesized and investigated for labeling those domains of the proteins which are outside the bilayer. For in vitro studies of phospholipid-protein interactions, a number of membrane proteins which are available in a pure state and possess well-defined functions will be studied. These include cytochrome b5, glycophorin A, beta-hydroxybutyrate dehydrogenase and Ca ions-ATPase of sarcoplasmic reticulum. Structural studies will include determination of sites of cross-linking and their variation as a function of the depth of the photosensitive group within the bilayer. Using the cross-linking approach for in vivo synthesis, incorporation of fatty acids containing the photosensitive groups into selected strains of E. coli and yeast will be further studied. Lipopolysaccharide of Gram-neagative bacteria is a powerful mitogen for B-lymphocytes whose structure has been recently elucidated in this laboratory. Systematic replacements of the fatty acids by photolabeled analogs will be carried out and the latter will be used in structure-function studies and in LPS-receptor isolation.
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