Mechanistic investigation of coordinated conformational changes in multisubunit ion channels and enzymes

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2008.Vita.Includes bibliographical references.Many enzymes and ion channels consist of multiple subunits and/or multiple distinct functional components. Coordinated conformational changes through allosteric interactions between subunits and/or between functional units can efficiently regulate protein activity. This dissertation describes investigations of coordinated conformational changes in two systems: the ATP-sensitive potassium (KATP) channel and the ATP-dependent bacterial protease, ClpAP. KATP channels consist of two protein subunits: a pore-forming subunit, Kir6.2 and a regulatory subunit, SUR1. Kir6.2 is an inwardly rectifying potassium channel, and SUR1 belongs to the ATP-binding cassette (ABC) superfamily. Using patch clamp techniques, KATP channel activity was observed directly with single-channel resolution. The results indicate that noise from stochastic channel gating is significantly reduced compared to what would be observed for identical and independent channels, and provide evidence that negatively cooperative interactions between neighboring KATP channels are the source of the noise reduction. Simulations further suggest that negative coupling among KATP channels in pancreatic beta cells could be important for reliable signal transduction. Energetic coupling between Kir6.2 and SUR1 subunits was also investigated. Single-channel records were analyzed to detect the violations of microscopic reversibility in channel gating that would occur if Kir6.2 conformational transitions were driven by the energy from ATP hydrolysis by SUR1. Although no violations of detailed balance in channel gating are detected on the time scale where ATP hydrolysis takes place, unexpected non-equilibrium gating is observed on longer time scales. These results imply that channel gating is coupled to non-equilibrium processes other than ATP hydrolysis by SUR1. The second system studied for coordinated conformational change was ClpAP.(cont.) ClpAP is composed of an ATPase, ClpA and a serine peptidase, ClpP. ClpA uses the free energy of ATP hydrolysis to unfold protein substrates and translocate them to ClpP, which proteolyzes them. To investigate how protein translocation by ClpA is coupled to proteolysis by ClpP, size distributions of peptide products were measured. The observation of non-exponential size distributions, in combination with simulations predicting how different mechanisms would influence the size distribution, supports the hypothesis that peptide product sizes are controlled by coordinated conformational changes of ClpA and ClpP.by Kee-Hyun Choi.Ph.D

Inelastic scattering in a monolayer graphene sheet; a weak-localization study

Charge carriers in a graphene sheet, a single layer of graphite, exhibit much distinctive characteristics to those in other two-dimensional electronic systems because of their chiral nature. In this report, we focus on the observation of weak localization in a graphene sheet exfoliated from a piece of natural graphite and nano-patterned into a Hall-bar geometry. Much stronger chiral-symmetry-breaking elastic intervalley scattering in our graphene sheet restores the conventional weak localization. The resulting carrier-density and temperature dependence of the phase coherence length reveal that the electron-electron interaction including a direct Coulomb interaction is the main inelastic scattering factor while electron-hole puddles enhance the inelastic scattering near the Dirac point.Comment: 12 pages, 3 figures, submitted to PR

Pseudogap Behavior Revealed in Interlayer Tunneling in Overdoped Bi2_2Sr2_2CaCu2_2O8+x_{8+x}

We report heating-compensated interlayer tunneling spectroscopy (ITS) performed on stacks of overdoped Bi$_2$Sr$_2$CaCu$_2$O$_{8+x}$ intrinsic junctions, where most of bias-induced heating in the ITS was eliminated. The onset temperature of the pseudogap (PG), revealed in the hump structure of the electronic excitation spectra, reached nearly room temperature for our overdoped intrinsic junctions, which represented the genuine PG onset. At a temperature below but close to $T_c$, both the superconducting coherence peak and the pseudogap hump coexisted, implying that the two gaps are of separate origins. The hump voltage increased below $T_c$, following the superconducting gap voltage, which led to a conclusion that the hump structure below $T_c$ in our ITS arose from the combined contribution of the quasiparticle spectral weights of two different characters; one of the superconducting state and another of the PG state near the antinodal region.Comment: accepted in Phys. Rev.

High Frequency Jet Ventilation of One Lung using a Bronchial Blocker of Univent during Carinal Resection

Airway management during carinal resection should provide adequate ventilation and oxygenation as well as a good surgical field, but without complications such as barotraumas or aspiration. One method of airway management is high frequency jet ventilation (HFJV) of one lung or both lungs. We describe a patient undergoing carinal resection, who was managed with HFJV of one lung, using a de-ballooned bronchial blocker of a Univent tube without cardiopulmonary compromise. HFJV of one lung using a bronchial blocker of a Univent tube is a simple and safe method which does not need additional catheters to perform HFJV and enables the position of the stiffer bronchial blocker more stable in airway when employed during carinal resection

WATCHFUL OBSERVATION VERSUS EARLY AORTIC VALVE REPLACEMENT FOR SYMPTOMATIC PATIENTS WITH LOW-GRADIENT SEVERE AORTIC STENOSIS AND PRESERVED EJECTION FRACTION

Brief Communications Arising: arising from X. Dong, B. Milholland & J. Vijg Nature 538, 257–259 (2016); doi:10.1038/nature19793. Comments by: Beer, J.A.A. de, Bardoutsos, A. & Janssen, F. (2017)

Health-Related Quality of Life Using the EuroQol 5D Questionnaire in Korean Patients with Type 2 Diabetes

We aimed; 1) to determine the validity of the EuroQol 5D (EQ-5D) for the health-related quality of life (HRQOL) of Korean patients with type 2 diabetes, and 2) to identify associated factors of the HRQOL of these patients. Follow-up surveys were conducted for consecutive patients with type 2 diabetes. HRQOL was assessed using the EQ-5D and the Short Form-36 (SF-36). The validity of EQ-5D was assessed with the perspectives of known group, convergent and discriminant validity. Additionally, a linear mixed model using a backward elimination was used for identify associated factors. Of the 1,072 patients included in the first survey, 858 (80.0%) completed the questionnaires in the follow-up. In the known group validity, the problem rates in each EQ-5D dimension were highest among women, elderly people, and less-educated subjects. The Spearman's ρ between the EQ-5D and the SF-36 scales were larger in the comparable dimensions than those in the less comparable dimensions. In the final model, we found that sex, age, education, body mass index, atrial fibrillation, stroke, and retinopathy were statistically significant. Our data suggest that the EQ-5D is a valid tool for Korean patients with type 2 diabetes and that various factors could affect their HRQOL