The cell physiology of addictive mechanisms involves seven- transmembrane receptors and their initiated signalling pathways. The phenomena of withdrawal and dependency have been correlated with the expression of cAMP signaling pathway components in several brain nuclei. Three primary elements play sequential roles in this pathway; receptor, G protein and adenylyl cyclase. I will test the hypothesis that mutations in the contact regions between proteins in this pathway can result in better coupling than wild-type. Bacterial screening mechanisms capable of selecting gain-of- function phenotypes from large, randomly mutated libraries will be used to identify mutations that augment binding between signaling molecules. This screening system has been constructed and previously implemented in this lab. Particular mutations that augment coupling will yield information about the structural characteristics of optimal adenylyl cyclase activating ligands, leading insight to therapeutic design. Bacterial without endogenous adenylyl cyclase cannot metabolize maltose. Transformation of bacteria with adenylyl cyclase and G protein has been shown to allow growth, with growth rate being dependent on the efficiency of coupling between the expressed molecules. A simple extension of this constructed pathway will include the expression of a G protein coupled, seven-transmembrane receptor in the inner-membrane of E. Coli, allowing for quick drug screening and efficient mutational analysis of the receptor.

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Individual Predoctoral NRSA for M.D./Ph.D. Fellowships (ADAMHA) (F30)
Project #
1F30DA005788-01
Application #
2012986
Study Section
Human Development Research Subcommittee (NIDA)
Project Start
1997-08-10
Project End
Budget Start
1997-02-10
Budget End
1998-02-09
Support Year
1
Fiscal Year
1997
Total Cost
Indirect Cost
Name
University of Chicago
Department
Biology
Type
Schools of Medicine
DUNS #
225410919
City
Chicago
State
IL
Country
United States
Zip Code
60637