Signal transduction by guanine nucleotide binding proteins (G proteins) plays a major role in the human cardiovascular, endocrine, and nervous systems. Many drugs used today including those for hypertension, angina, and congestive heart failure have actions on G protein coupled receptors. Excitingly, recent studies show that these receptors can activate G protein cascades in the absence of their normal physiological stimulus. Mutations leading to """"""""runaway receptors"""""""" are the basis for several rare genetic disorders. Similar mechanisms may contribute to more common polygenic disorders (such as hypertension, diabetes, and heart failure) which are epidemic in our society today. Drugs which block receptor activation by physiological stimuli would not be expected to inhibit a """"""""runaway receptor"""""""". A major goal of this project is to understand the structural basis of the G protein signalling events immediately downstream of the receptor with the goal of designing drugs to block at those sites and effectively inhibit all receptor function. Thus, the PI will develop a functional map of G protein sites which contact receptors and effectors. G proteins are composed of three subunits, alpha, beta, and gamma. Much is known about alpha subunits, but much less is known about the beta and gamma subunits which have only recently been recognized to play an active part in signalling.
Specific aims of this project are: 1) Receptor-G protein interface: The PI will define the roles of intracellular regions of alpha2-adrenergic and angiotensin II receptors in coupling to different G proteins (Gi, Gs, and Gq). Identifying specificity determinants and activation determinants will help in the design of targetted G protein activators and inhibitors. Combinatorial peptide libraries of receptor fragments will be used to discover peptides with high affinity and specificity for G proteins. Such peptides will provide new knowledge about receptor-G protein interfaces and will represent useful structural leads for drug design. 2) Beta subunit structural map: The PI recently provided the first evidence about which region of the G-beta subunit binds to receptors. The PI will refine the localization of this receptor-beta subunit contact site and define the loci of beta-gamma/effector interactions with a combined approach using biochemical labelling, synthetic peptides, and genetic selection in yeast. 3) Delivery and targetting of G protein inhibitors: To facilitate delivery of peptide drugs as potential therapeutic agents, the PI will utilize DNA-mediated delivery of receptor and G protein fragments (minigenes). The specificity of these """"""""drugs"""""""" will be tested in cell culture with the aim of future use in gene therapy approaches to cardiovascular disease.
Showing the most recent 10 out of 25 publications
