We propose to continue our investigation of red cell membrane proteins with emphasis on those components directly or indirectly associated with intramembrane particles. With our recently developed fusion system for studying the migration of membrane components into newly fused large-sized liposome regions, we will analyze the structural relationships between intramembrane particles and other components in the native membrane. Intramembrane particles are detected by freeze-fracture electron microscope whereas migration of individual proteins are identified by ferritin-tagged antibodies and/or lectins. Such approaches have so far provided evidence for the existence of a control mechanism by cytoskeletal proteins for intramembrane particle migration. We will now extend these studies to delineate the particular components responsible for such restraint by means of fusion between liposomes and membrane preparations containing different combinations of membrane components and also by the use of reagents perturbing interactions among these components. The major component of intramembrane particles, Band 3, has been implicated as the site of anion transport, a crucial physiological function in red cells which enables the transfer of CO2. We propose to prepare monoclonal hybridoma antibodies selected by their ability to bind to Band 3, to compete with the binding of transport inhibitor and to modulate anion transport. These homogeneous antibodies with exquisite specificity have the discriminatory power to identify novel antigenic determinants involved in anion transport on either Band 3 or other membrane components of RBC membrane. They can also be used to dissect the functional segments of a given transport protein and to analyze the molecular basis of this putative transport activity in intramembrane particles.
| Matsumoto, S S; Yu, A L; Yu, J (1985) Morphological changes in leukemic lymphoblasts and normal lymphocytes treated with deoxyadenosine plus deoxycoformycin. Cancer Invest 3:225-33 |
