A long-pursued goal of structural biology has been the determination of the structure of membrane proteins in their native membrane environment, and in their various functional states as defined by transmembrane gradients of substrate molecules and membrane potentials. Application of such gradients, let alone crystallization in a membrane environment, has not been possible in the case of X-ray structure determination. The application of gradients has remained out of reach as well for the analysis of 2D crystals by electron crystallography. The proposed work concerns the further development of a general method for single-particle cryo-EM structure determination of membrane proteins reconstituted into liposomes. The method is called random spherically- constrained (RSC) single-particle reconstruction. In the proposed work we will work to surmount several remaining technical hurdles, and then apply this combination of biochemistry, EM imaging and image processing methods toward a nanometer- resolution structure of the GluA2 glutamate receptor as reconstituted into a lipid membrane.

Public Health Relevance

In order to understand the detailed workings of the molecular machines that underlie life, one needs to examine their three-dimensional structures. In the proposed work we will work to surmount several technical hurdles to obtain 3D structures of GluA2 glutamate receptor in a membrane environment. This glutamate receptor is an essential signal- reception machine in brain cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21NS085776-01
Application #
8635183
Study Section
Special Emphasis Panel (ZRG1-MDCN-J (04))
Program Officer
Silberberg, Shai D
Project Start
2013-09-01
Project End
2015-08-31
Budget Start
2013-09-01
Budget End
2014-08-31
Support Year
1
Fiscal Year
2013
Total Cost
$246,425
Indirect Cost
$96,425
Name
Yale University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520