William Gullick

STRUCTURE AND FUNCTION OF THE ACETYLCHOLINE RECEPTOR MOLECULE STUDIED USING MONOCLONAL ANTIBODIES

Acetylcholine receptors ( AChRs) are the object of an antibody-mediated autoimmune response in myasthenia gravis (MG) and its animal model, experimental autoimmune myasthenia gravis (EAMG) (reviewed in references 1-4). In order to understand how AChR might provoke an immune response or respond to an immune attack, it is important to understand the structure and function of the AChR molecule. And, in order to understand how antibodies to the AChR act, it is important to determine what specificities of antibodies can be made and what effect they have on the AChR. The structure and function of the AChR are also of fundamental interest, because it is the archetypic neurotransmitter receptor and a model surface membrane protein. Antisera containing heterogeneous populations of antibodies are useful reagents, but pure antibodies of each type would be better. When an animal is immunized with AChR it makes a large population of antibodies directed at many determinants on the molecule. As a result of differences in their “variable” regions, these antibodies differ in which antigenic determinants they bind and in the affinity with which they bind to these determinants; and as a result of differences in their “constant” regions, these antibodies differ in their ability to interact with other immune effectors like complement and phagocytes. Several aspects of AChR structure, function and the pathology of MG could be very effectively studied using homogeneous antibody reagents. For example, most antibodies bind to AChRs without affecting the acetylcholine binding sites or the cation channel they reg~late.~--” Such antibodies, if obtained as individual reagents, would be excellent probes for structures on the AChR macromolecule other than the acetylcholine binding site. The acetylcholine binding site comprises only a tiny part of the extracellular surface of this large macromolecule, and is in no great need of probes, since snake venom toxins,1° affinity labeling reagents,11 and reversible agonists and antagonists are available in abundance. Some antibodies in anti-AChR sera can effect the length of time that the cation channel triggered by acetylcholine is open or its conductance when open.?, 12 Such antibodies, if obtained as individual reagents, would be excellent probes for the AChR channel, for which few other probes exist. LOSS

IMMUNOCHEMICAL CHARACTERIZATION OF ACETYLCHOLINE RECEPTORS USING MONOCLONAL ANTIBODIES

By two different competitive binding techniques using intact AChR, the same set of at least six antigenic determinants or regions were distinguished in the a subunits., Most mAbs and most antibodies in antisera were directed at a main immuqogenic region (MIR) on a. Those mAbs which also reacted with the denatured a subunit bound characteristic patterns of proteolytic fragments of a subunit corresponding to four of the regions, including the MIR. This grouping was also reflected by the binding capacity of these mAbs to the membrane bound AChR.3