The long-term aim of this work is to understand the structural components of the BK-type calcium (Ca2+)-activated potassium (K+) channel complex, the functional properties of the channels, and the physiological roles of those channels. BK channels are widely expressed among different cell types and exhibit significant functional diversity suited to their physiological roles. BK currents couple changes in submembrane Ca2+ concentrations to changes in membrane potential and excitability. They consist minimally of four pore-forming subunits. In addition, there are four auxiliary subunits, which differ not only in terms of tissue distribution, but also in terms of the functional properties of the resulting BK channels. Two auxiliary subunits, the 2 and 3b, produce a novel rapid inactivation of BK channel current. This project will focus specifically on the mechanism and structural basis of inactivation mediated by these subunits. Inactivation resulting from these subunits involves a two-step mechanism unlike any inactivation mechanism so far described. Using methods of electrophysiology combined with molecular biology, this project will determine the structural components and molecular steps involved in the inactivation mechanism produced by the 2 and 3b subunits. First, a model by which inactivation domains bind to a pore constriction at the mouth of the channel will be functionally tested. Second, the points of interaction between inactivation domains and their binding sites will be determined. Third, the validity of two-step inactivation mechanism will be confirmed and the molecular nature of each of the two steps in the inactivation process determined. These studies will provide important information about key structural changes associated with BK channel gating and the structural components that contribute to the channel near the channel pore. BK channels are of broad importance in the normal functioning of a variety of excitable cells. Among different tissues, BK channels contribute to regulation of neuronal excitability, smooth muscle relaxation, synaptic transmission and hormone release. Better understanding the composition and functional role of BK channel variants is of potential medical importance, not only because the channels may serve as specific therapeutic targets but also because altered function of particular variants may contribute to unknown pathological conditions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK046564-11
Application #
6846625
Study Section
Special Emphasis Panel (ZRG1-MDCN-4 (01))
Program Officer
Jones, Teresa L Z
Project Start
1993-05-21
Project End
2007-01-31
Budget Start
2005-02-01
Budget End
2006-01-31
Support Year
11
Fiscal Year
2005
Total Cost
$336,600
Indirect Cost
Name
Washington University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Zeng, Xu-Hui; Benzinger, G Richard; Xia, Xiao-Ming et al. (2007) BK channels with beta3a subunits generate use-dependent slow afterhyperpolarizing currents by an inactivation-coupled mechanism. J Neurosci 27:4707-15
Zhang, Zhe; Zhou, Yu; Ding, Jiu-Ping et al. (2006) A limited access compartment between the pore domain and cytosolic domain of the BK channel. J Neurosci 26:11833-43
Benzinger, G Richard; Xia, Xiao-Ming; Lingle, Christopher J (2006) Direct observation of a preinactivated, open state in BK channels with beta2 subunits. J Gen Physiol 127:119-31
Chen, Xiao-Ke; Wang, Lie-Cheng; Zhou, Yang et al. (2005) Activation of GPCRs modulates quantal size in chromaffin cells through G(betagamma) and PKC. Nat Neurosci 8:1160-8
Xia, Xiao-Ming; Ding, J P; Lingle, Christopher J (2003) Inactivation of BK channels by the NH2 terminus of the beta2 auxiliary subunit: an essential role of a terminal peptide segment of three hydrophobic residues. J Gen Physiol 121:125-48
Wang, Ying-Wei; Ding, Jiu Ping; Xia, Xiao-Ming et al. (2002) Consequences of the stoichiometry of Slo1 alpha and auxiliary beta subunits on functional properties of large-conductance Ca2+-activated K+ channels. J Neurosci 22:1550-61
Ding, Jiu Ping; Lingle, Christopher J (2002) Steady-state and closed-state inactivation properties of inactivating BK channels. Biophys J 82:2448-65
Zhang, X; Solaro, C R; Lingle, C J (2001) Allosteric regulation of BK channel gating by Ca(2+) and Mg(2+) through a nonselective, low affinity divalent cation site. J Gen Physiol 118:607-36
Zeng, X H; Ding, J P; Xia, X M et al. (2001) Gating properties conferred on BK channels by the beta3b auxiliary subunit in the absence of its NH(2)- and COOH termini. J Gen Physiol 117:607-28
Lingle, C J; Zeng, X H; Ding, J P et al. (2001) Inactivation of BK channels mediated by the NH(2) terminus of the beta3b auxiliary subunit involves a two-step mechanism: possible separation of binding and blockade. J Gen Physiol 117:583-606

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