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TSE COUNCIL FOR TOBACCO RESEARCH-U.S.A., INC. Memorandum To: Dr. H.McAllister and Staff From: D.H.Ford Re: Site visit with Dr.Gary Yellen, Neuroscience Center, MGH East, July 30, 1992. Site Visitors D.H.Ford and A.Eisman. Grant No. 3243 entitled "Mutagenesis studies of the activation site of the nicotinic acetylcholine receptor." Goals: To learn more concerning the structure and function of the nACh ligand binding site by: 1. learning the role of specific amino acid residues in the binding and activation of the receptor. 2.To determine the key residues for ligand binding by site-directed mutagenesis and phenotypic seclection of nAChR mutants which are specifically altered in their affinity or response to specific ligands. These are extremely ambitious goals which have become somewhat more difficult to ach eve in the last few months inasmuch as it is now known that there are now 8 versions of the alpha peptide and 6 versions of the beta peptide which contribute to the receptor. Comment:Dr. Yellen initiated his move from Johns Hopkins in January and was able to have some of his people undertake some of the work described in his proposal. However, the task of -:estanlishing his laboratory is still underway and thus his program is not yet in full swing. Now he understands that he may be required to move to other facilities at Harvard in the Dept. of Neurobiology. While there is a plus to this move in that he will be near colleagues with whom he collaborates, the move will again disrupt his research program and delay his progress. To return to his goals: The concept that there are now 8 versions of the alpha peptid'e, to which ligand binding occurs and 6 versions of the beta peptide, which is in someway involved with the structural integrity of the receptor leads to,an interesting array of concepts which need to be proven. 1. Do changes in the alpha peptide amino acid composition influence the binding of various ligands and the affinity of binding? Similarly, does an alteration in a struture dependent peptide influence receptor binding? Do these variants occur at the neuromuscular junction as much as in the CNS where 8 versions of the receptor have already been detected? Clearly, with the number of ligand binding peptides and structural peptide variants present in the nAChR, there may have to be a great many mutants created to determine which are the key residues, since this may vary with each receptor subtype. This is a very interesting program, which because of having to move his laboratory has caused Dr. Yellen to progress more slowly that he had anticipated. It is also of importance if one is ever to understand apparent variations in response of subtypes of CNS nACh receptors (See Lorna Role), which may be related to behavioural differences as well...or differences in response of various types of neurons,ie. stimulation of synthesis and release of dopamine and inhibition of release of norepinephrine. DHF