This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Ion-conductive channels are membrane proteins playing a vital regulatory role in all living organisms. Their failure to function properly results in a variety of diseases, such as neurological disorders. An essential function of channels is gating, activated by a variety of stimuli and accompanied by substantial structural changes. The X-ray data on 2-TM KcsA and MthK K+ channels led to a model based on a hinge-bending mechanism of gating. The 30? bending about the conserved Gly99 residue of the TM2-domain in MthK results in wide splay opening of the intracellular gating pore. KcsA has glycine 83 at the similar position of TM2. The model, however, contradicted an earlier study by cw-ESR that suggested smaller TM2 motion. Very recent data on gating in the Shaker channel has shown that the open conformation of MthK may not be applicable to other K+ channels. Thus, knowing the structure of KcsA in its open state is of utmost importance in our understanding of the mechanism of gating in K+ channels. Therefore, our current efforts in collaboration with Perozo are centered on the study of the pH induced global conformation rearrangement of KcsA using fully-labeled channels, tandem dimers, and tandem tetramers. This will provide both inter- and intra-domain distances to elucidate the global structural changes. Large structural changes require measuring distances of up to 50 . In our preliminary study on KcsA and MscL we have already demonstrated the power of DQC-ESR. We successfully applied DQC to WT KcsA fully-labeled at C28, and tandem dimer constructs, labeled at 87 and 119. For spin-labels in contact with lipids short phase relaxation times limits accurate measurements to ca. 35 . Therefore, in the cases when distances are large we will use lipids with perdeuterated acyl chains (i.e. DMPC-d54), which let us accurately measure distance up to 50 .

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR016292-07
Application #
7602580
Study Section
Special Emphasis Panel (ZRG1-BCMB-K (40))
Project Start
2007-09-01
Project End
2008-08-31
Budget Start
2007-09-01
Budget End
2008-08-31
Support Year
7
Fiscal Year
2007
Total Cost
$961
Indirect Cost
Name
Cornell University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
872612445
City
Ithaca
State
NY
Country
United States
Zip Code
14850
Jain, Rinku; Vanamee, Eva S; Dzikovski, Boris G et al. (2014) An iron-sulfur cluster in the polymerase domain of yeast DNA polymerase ?. J Mol Biol 426:301-8
Pratt, Ashley J; Shin, David S; Merz, Gregory E et al. (2014) Aggregation propensities of superoxide dismutase G93 hotspot mutants mirror ALS clinical phenotypes. Proc Natl Acad Sci U S A 111:E4568-76
Georgieva, Elka R; Borbat, Peter P; Ginter, Christopher et al. (2013) Conformational ensemble of the sodium-coupled aspartate transporter. Nat Struct Mol Biol 20:215-21
Airola, Michael V; Sukomon, Nattakan; Samanta, Dipanjan et al. (2013) HAMP domain conformers that propagate opposite signals in bacterial chemoreceptors. PLoS Biol 11:e1001479
Airola, Michael V; Huh, Doowon; Sukomon, Nattakan et al. (2013) Architecture of the soluble receptor Aer2 indicates an in-line mechanism for PAS and HAMP domain signaling. J Mol Biol 425:886-901
Yang, Zhongyu; Liu, Yangping; Borbat, Peter et al. (2012) Pulsed ESR dipolar spectroscopy for distance measurements in immobilized spin labeled proteins in liquid solution. J Am Chem Soc 134:9950-2
Yu, Long-Xi; Dzikovski, Boris G; Freed, Jack H (2012) A protocol for detecting and scavenging gas-phase free radicals in mainstream cigarette smoke. J Vis Exp :e3406
Sun, Yan; Zhang, Ziwei; Grigoryants, Vladimir M et al. (2012) The internal dynamics of mini c TAR DNA probed by electron paramagnetic resonance of nitroxide spin-labels at the lower stem, the loop, and the bulge. Biochemistry 51:8530-41
Smith, Andrew K; Freed, Jack H (2012) Dynamics and ordering of lipid spin-labels along the coexistence curve of two membrane phases: an ESR study. Chem Phys Lipids 165:348-61
Yu, Renyuan Pony; Darmon, Jonathan M; Hoyt, Jordan M et al. (2012) High-Activity Iron Catalysts for the Hydrogenation of Hindered, Unfunctionalized Alkenes. ACS Catal 2:1760-1764

Showing the most recent 10 out of 72 publications