9419175 McLaughlin The overall objective is to understand the molecular mechanism by which proteins containing myristate, a 14 carbon acyl chain, interact with biological membranes. The proposal focuses on Src (the product of the v-src oncogene, v-Src, and its cellular homolog, c-Src, are known collectively as Src); the working hypothesis is that membrane binding requires both the hydrophobic insertion of the NH2-terminal myristate into the bilayer and the electrostatic interaction of a cluster of adjacent basic residues with acidic lipids. There are four specific projects. 1) Developing a simple model that describes the synergistic effect of myristate and basic residues on the binding of Src to phospholipid membranes and testing this model by measuring the membrane binding of myristoylated and nonmyristoylated peptides corresponding to the NH2-terminal region of Src. 2) Confirming that the basic residues in the NH2-terminal region of Src are responsible for the electrostatic component of protein-membrane interaction by measuring the membrane binding of native v-Src and c-Src as well as mutant and chimeric versions of these proteins. 3) Measuring the effect of phosphorylation by C and A kinases on the membrane binding of both Src and corresponding peptides. 4) Constructing a molecular model of a phospholipid bilayer and the NH2-terminal region of Src, then using the nonlinear Poisson-Boltzmann equation to calculate theoretically the electrostatic component of the binding. The project is significant because membrane binding is crucial to the function of Src. The biophysical principles that emerge should be applicable to some other myristoylated (e.g. MARCKS) and farnesylated (e.g. K-Ras) proteins. %%% The overall objective is to understand the molecular mechanism by which proteins containing myristate, a 14 carbon acyl chain, interact with biological membranes. The proposal focuses on Src (the product of the v-src oncogene, v-Src, which functions to transfor m cells, and its cellular homolog, c-Src, which is involved in control of the cell cycle, are known collectively as Src); the working hypothesis is that membrane binding requires both the hydrophobic insertion of the NH2-terminal myristate into the bilayer and the electrostatic interaction of a cluster of adjacent basic residues with acidic lipids. These are four specific projects. 1) Developing a simple model that describes the synergistic effect of myristate and basic residues on the binding of Src to phospholipid membranes and testing this model by measuring the membrane binding of synthetic peptides. 2) Confirming that the basic residues in the NH2-terminal region of Src are responsible for the electrostatic component of protein-membrane interaction by measuring the membrane binding of native v-Src and c-Src as well as mutant and chimeric versions of these proteins. 3) Measuring the effect of phosphorylation by C and A kinases on the membrane binding of both Src and corresponding peptides. 4) Constructing a molecular model of a phospholipid bilayer and the NH2-terminal region of Src, then calculating theoretically the electrostatic component of the binding. The project is significant because membrane binding is crucial to the function of Src and the biophysical principles that emerge should be applicable to other myristoylated proteins. ***
