Abstract

Since Anfinsen's demonstration that the sequence of a protein dictates its structure, the energetics and kinetics of folding have been explored with ever-greater temporal and spatial resolution. However, almost all of these studies have been devoted to water-soluble proteins, and our understanding of membrane proteins is in its infancy. Here, we use protein design to test and refine our understanding of membrane protein folding. Model peptides are designed to associate into helical bundles in micelles and phospholipid bilayers. The study of these systems is simplified by the fact that the unfolded state is well defined (the monomeric helices), and the energetics of folding can be monitored by measuring the monomer-nmer equilibrium.
In aims 1 & 2, we test the importance and relative energetic contributions of disparate forces such as van der Waals packing versus polar interactions. These studies will not only further our understanding of protein folding, but also provide insight and methods to understand the process of transmembrane helix association, which often plays an essential role in signal transduction. As an outgrowth of our understanding of the folding of membrane proteins, we are beginning to design water-soluble versions of 2 membrane proteins, phospholamban and the KcsA potassium channel. These studies promise to provide a new method for obtaining large quantities of water-soluble versions of membrane proteins for pharmaceutical and biophysical studies.
Our specific aims are as follows:
Aim 1. We will study the association of designed and natural transmembrane peptides, and use these systems to understand the features leading to the folding and association of transmembrane helices.
Aim 2. We will explore how regions of a protein located in the aqueous, headgroup, and membrane interior cooperate to dictate the fold of a protein by designing helical bundles that incorporate both transmembrane as well as water-soluble folding motifs.
Aim 3 We will structurally characterize water-soluble versions of phospholamban and KcsA.
Aim4. We will develop computational methods for transmembrane protein structure prediction & design. ? ?

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM060610-06
Application #
7233254
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Chin, Jean
Project Start
1999-12-01
Project End
2010-04-30
Budget Start
2007-05-01
Budget End
2008-04-30
Support Year
6
Fiscal Year
2007
Total Cost
$278,694
Indirect Cost

  Institution

Name
University of Pennsylvania
Department
Biochemistry
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104

  Publications

Soto, Cinque S; Hannigan, Brett T; DeGrado, William F (2011) A photon-free approach to transmembrane protein structure determination. J Mol Biol 414:596-610
Remorino, Amanda; Korendovych, Ivan V; Wu, Yibing et al. (2011) Residue-specific vibrational echoes yield 3D structures of a transmembrane helix dimer. Science 332:1206-9
Shandler, Scott J; Korendovych, Ivan V; Moore, David T et al. (2011) Computational design of a ?-peptide that targets transmembrane helices. J Am Chem Soc 133:12378-81
Zhu, Hua; Metcalf, Douglas G; Streu, Craig N et al. (2010) Specificity for homooligomer versus heterooligomer formation in integrin transmembrane helices. J Mol Biol 401:882-91
Goldberg, Shalom D; Clinthorne, Graham D; Goulian, Mark et al. (2010) Transmembrane polar interactions are required for signaling in the Escherichia coli sensor kinase PhoQ. Proc Natl Acad Sci U S A 107:8141-6
Berger, Bryan W; Kulp, Daniel W; Span, Lisa M et al. (2010) Consensus motif for integrin transmembrane helix association. Proc Natl Acad Sci U S A 107:703-8
Zhang, Yao; Kulp, Daniel W; Lear, James D et al. (2009) Experimental and computational evaluation of forces directing the association of transmembrane helices. J Am Chem Soc 131:11341-3
Weeks, Colin L; Polishchuk, Alexei; Getahun, Zelleka et al. (2008) Investigation of an unnatural amino acid for use as a resonance Raman probe: Detection limits, solvent and temperature dependence of the nuC identical withN band of 4-cyanophenylalanine. J Raman Spectrosc 39:1606-1613
Caputo, Gregory A; Litvinov, Rustem I; Li, Wei et al. (2008) Computationally designed peptide inhibitors of protein-protein interactions in membranes. Biochemistry 47:8600-6
Grigoryan, Gevorg; Degrado, William F (2008) Modest membrane hydrogen bonds deliver rich results. Nat Chem Biol 4:393-4

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