Voltage gated proton channels are proteins found in many cell membranes that allow protons to cross the membrane in a highly regulated manner. For over two decades, these molecules were studied in cells using electrical measurements (voltage-clamp). The identification in 2006 of the gene that codes for these molecules makes it possible to study the molecular structure in unprecedented detail. In this project, specific mutations in the proton channel gene will be produced and the mutant channels will be expressed in cell lines. The effects these mutations have on molecular behavior will be evaluated using patch-clamp measurements. One goal is to determine how proton channels sense pHo and pHi, the proton concentrations on either side of the cell membrane, which is crucial to their ability to carry out their functions. We hypothesize that one or a few amino acid residues that are accessible to the solution bind protons and thus act as pH sensors. The second goal is to determine which parts of the molecule sense membrane potential. Because the proton channel molecule resembles the voltage-sensing domains of other ion channels, the first attempt will be to determine if the same residues sense voltage. Next, other charged amino acids located in membrane-spanning regions will be examined. Third, the location of the pathway by which protons pass through the molecule will be determined. The first hypothesis tested will be based on similarities to corresponding domains of other channels. The unique aspect of proton channels is a conduction mechanism of proton hopping (Grotthuss-like) that could occur at a titratable reside. Next the accessibility to external or internal solutions of amino acid residues at specific locations in the molecule will be examined systematically, by inserting cysteine residues at specific locations and then determining the rate at which they can be modified chemically ("cysteine scanning"). Finally, the discovery in 2008 that proton channel is a homodimer raises questions that will be addressed. We will test whether the inhibitor Zn2+ binds with high affinity between the two monomers. These and other experiments will indicate whether the two monomers function independently, or if they interact.

Public Health Relevance

Proton channels are proteins that white blood cells need in order to kill bacteria and other microbial invaders. This project will identify the working parts of the proton channel molecule so that drugs to regulate proton channel function can be developed.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM087507-04
Application #
8460040
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Chin, Jean
Project Start
2010-05-01
Project End
2014-04-30
Budget Start
2013-05-01
Budget End
2014-04-30
Support Year
4
Fiscal Year
2013
Total Cost
$279,440
Indirect Cost
$93,147
Name
Rush University Medical Center
Department
Physiology
Type
Schools of Medicine
DUNS #
068610245
City
Chicago
State
IL
Country
United States
Zip Code
60612
DeCoursey, Thomas E; Hosler, Jonathan (2014) Philosophy of voltage-gated proton channels. J R Soc Interface 11:20130799
Morgan, Deri; Decoursey, Thomas E (2014) Analysis of electrophysiological properties and responses of neutrophils. Methods Mol Biol 1124:121-58
Kulleperuma, Kethika; Smith, Susan M E; Morgan, Deri et al. (2013) Construction and validation of a homology model of the human voltage-gated proton channel hHV1. J Gen Physiol 141:445-65
DeCoursey, Thomas E (2013) Voltage-gated proton channels: molecular biology, physiology, and pathophysiology of the H(V) family. Physiol Rev 93:599-652
Smith, Susan M E; DeCoursey, Thomas E (2013) Consequences of dimerization of the voltage-gated proton channel. Prog Mol Biol Transl Sci 117:335-60
Musset, Boris; Cherny, Vladimir V; DeCoursey, Thomas E (2012) Strong glucose dependence of electron current in human monocytes. Am J Physiol Cell Physiol 302:C286-95
Musset, Boris; Clark, Robert A; DeCoursey, Thomas E et al. (2012) NOX5 in human spermatozoa: expression, function, and regulation. J Biol Chem 287:9376-88
Smith, Susan M E; Morgan, Deri; Musset, Boris et al. (2011) Voltage-gated proton channel in a dinoflagellate. Proc Natl Acad Sci U S A 108:18162-7
Musset, Boris; Smith, Susan M E; Rajan, Sindhu et al. (2011) Aspartate?112 is the selectivity filter of the human voltage-gated proton channel. Nature 480:273-7
Capasso, Melania; DeCoursey, Thomas E; Dyer, Martin J S (2011) pH regulation and beyond: unanticipated functions for the voltage-gated proton channel, HVCN1. Trends Cell Biol 21:20-8

Showing the most recent 10 out of 15 publications