The activation of adenylyl cyclase by B adrenergic receptors (BAR) will be investigated using complete membrane systems from S49 mouse lymphoma and other cells. The kinetic and thermodynamic interactions of receptors, G proteins and the catalytic unit will be deduced using mathematical models. The overall specific aim is to be able to predict the consequence on adenylyl cyclase activity of any change in the concentration of receptor, G protein or catalytic unit or of any change in their kinetic or thermodynamic properties. Such changes inevitably accompany desensitization of the cell by drugs and are the vehicle by which phenotypically significant mutations are expressed. These studies make extensive use of mathematical kinetic models which relate betaAR concentration, G protein concentration and drug type to adenylyl cyclase activity. Recent developments in the thermodynamics of the components allow the drug induced changes in affinities of receptors for G proteins to be assessed more precisely than was previously possible. Hypotheses are proposed to relate the kinetic models to fundamental thermodynamic measurements. These hypotheses will be tested. The betaAR/Gs/adenylyl cyclase system is the best described and understood of the receptor G protein effector systems. It is to be expected that a knowledge of the mechanism of adenylyl cyclase activation will have wide applicability in understanding the control of G protein activity in general. Methodology includes a) ligand binding kinetics, b) mathematical modeling c) enzymology with adenylyl cyclase, and d) recombinant DNA technology. It is expected that a quantitative knowledge of adenylyl cyclase activation should lead to a better understanding of drug induced desensitization (tolerance) and of tumors associated with mutant G proteins, and to improved drug design for the clinically important B adrenergic agonists.
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