The movements that occur during protein functional cycles often involve twists and bends of secondary structure elements. In the case of transmembrane helices, the mechanisms for directing these conformational changes are likely to be particularly important because breaking and stretching the strong backbone hydrogen bonds is not energetically trivial. Thus, to understand membrane protein conformational changes, we need to learn how backbone movements are encoded by the sequence. We propose to scrutinize the mechanisms of transmembrane helix distortion both in an isolated helix and in a full length membrane protein through a battery of experimental methods, coupled to detailed molecular dynamics simulations.
The specific aims are:
Aim 1. How do sequence changes alter H-bond shifting, H-bond strengths and dynamics within an isolated model TM helix? Starting with a TM helix ?host?, we will introduce ?guest? amino acids and measure (1) alterations in i, i+4 to i, i+3 H-bond shifts by NMR methods; (2) alterations in helical dynamics by EPR methods; and (3) alterations in backbone H-bond strengths by measuring H/D isotope effects. To obtain a mechanistic understanding, we will study the sequence effects via molecular dynamics (MD) simulations. This work will illuminate how backbone dynamics can be encoded in a TM helix.
Aim 2. How do alterations in backbone H-bonding and dynamics affect function? To test the importance of TM helix flexibility in conformational signal transduction through a helix, we will study the KvAP channel. We will introduce backbone ?ester? mutations and side chain mutations to alter backbone hydrogen bonding and flexibility. Alterations in backbone dynamics and hydrogen bonding will be validated experimentally and the effects on voltage sensing investigated. Detailed visualization of the experimentally observed changes will be explored via MD simulations.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Specialized Center--Cooperative Agreements (U54)
Project #
5U54GM087519-08
Application #
9351550
Study Section
Special Emphasis Panel (ZGM1)
Project Start
2010-08-10
Project End
2019-08-31
Budget Start
2017-09-01
Budget End
2018-08-31
Support Year
8
Fiscal Year
2017
Total Cost
Indirect Cost
Name
University of Chicago
Department
Type
DUNS #
005421136
City
Chicago
State
IL
Country
United States
Zip Code
60637
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