Studies of Allostery in the Potassium Channel Kcsa by Solid-state NMR

In this thesis, I focus on studies of the mechanism of inactivation in KcsA. Allosteric coupling between the pH gate and the selectivity filter in the protein is hypothesized to be the cause of inactivation. Allosteric coupling refers to changes at one site of a protein due to perturbations at a remote site. In chapter 3, I measured the potassium affinities at the selectivity filter at neutral and low pH, which corresponds to the closed and open conformation at the pH gate. The results show a three order of magnitude shift in the potassium affinity. This is direct evidence that the pH gate and the selectivity filter are coupled, in support of the activation-coupled inactivation hypothesis. The allosteric coupling factor, defined as the ratio of the affinities, can be used as a benchmark to study other factors in the allosteric process, such as the membrane and specific residues. Because of the potential deleterious effect of the acidic pH on the protein and membrane, we studied a mutant E118A&H25R, in which the pH gate is mutated to be open. Thus we were able to measure the K+ affinity change in the open and closed conformation at the pH gate at neutral pH. The results confirmed that the opening of the pH gate results in an energetic stabilization of the collapsed (K+-unbound) state, and shifts the K+ affinity towards looser binding. In chapter 4, I tested the important role of residue F103 in mediating allosteric coupling, as suggested by electrophysiology and crystallography studies. I mutated this residue and measured the allosteric coupling factor on the mutant. The affinity at low pH is much tighter than wild-type and the coupling factor is significantly reduced. From the spectra, I observe local structural changes on I100 and T74 as a result of F103A mutation, implying the interaction among F103, I100 and T74 to mediate the allosteric coupling. F103 is distant from the pH gate and the selectivity filter; its effect on the coupling and inactivation behaviors confirms that inactivation involves coupling between the pH gate and the selectivity filter. In chapter 5, I developed a method to probe those allosteric participants, such as F103 in KcsA by NMR measurements. I tested this method on KcsA, dissecting KcsA into various functional compartments. Various allosteric participants T75Cg T74Cg I100 were identified. The importance of residue T74 for the coupling was confirmed by electrophysiology and NMR thermodynamics characterization. In chapter 6, we applied SSNMR to probe the structural and magnetic properties of superatom clusters

Deqi Sensations of Transcutaneous Electrical Nerve Stimulation on Auricular Points

Deqi sensation, a psychophysical response characterized by a spectrum of different needling sensations, is essential for Chinese acupuncture clinical efficacy. Previous research works have investigated the component of Deqi response upon acupuncture on acupoints on the trunk and limbs. However, the characteristics of Deqi sensations of transcutaneous electrical nerve stimulation (TENS) on auricular points are seldom reported. In this study, we investigated the individual components of Deqi during TENS on auricular concha area and the superior scapha using quantitative measurements in the healthy subjects and depression patients. The most striking characteristics of Deqi sensations upon TENS on auricular points were tingling, numbness, and fullness. The frequencies of pressure, warmness, heaviness, and soreness were relatively lower. The dull pain and coolness are rare. The characteristics of Deqi were similar for the TENS on concha and on the superior scapha

The homo- and hetero-oligomeric foldomes of functional and pathological amyloids

17th Iberian Peptide Meeting, Madrid, 5-7th February 2020This work was supported by grants from the NIH (R01 AI045937 to H.W and A.E.M.) and La Caixa Foundation (Junior Leader Project LCF/BQ/PR19/11700003 to MM). NMR experiments were performed at the “Manuel Rico” NMR Laboratory (LMR) of the Spanish National Research Council (CSIC), and at the New York Structural Biology Center (NYSBC). The LMR is a node of the Spanish Large-Scale National Facility (ICTS-R-LRB). The NYSBC is a STAR center supported by the New York State Office of Sciecne, Technology and Academic Research.Peer reviewe

Investigation into climate change effects on carbon and water fluxes, and water use efficiency of the temperate grassland ecosystems in Inner Mongolia of China

The daily gross primary productivity (GPP) and evapotranspiration (ET) in the Xilingol grassland ecosystem of Inner Mongolia were simulated using the BioGeochemical Cycles (Biome-BGC) model for 2003–2019 and under future climate-change scenarios. The system was optimized using the System Response Parameter Calibration Method (SRPCM). The temporal variations of GPP, ET and water use efficiency (WUE) were investigated, and the impacts of precipitation and temperature were explored. Results showed that (i) the BIOME-BGC model performed better when optimized using the SRPCM than by applying the Model-Independent Parameter Estimation approach (PEST); (ii) GPP and ET at annual and seasonal scales showed an insignificant increasing trend; (iii) WUE at the annual scale and in growing seasons showed an insignificant increasing trend and a slight decreasing trend in non-growing seasons; (iv) annual GPP and ET were more sensitive to changes in precipitation than changes in temperature with WUE keeping relatively stable with years; (v) precipitation is a critically controlling factor for GPP and ET in growing seasons and for ET and WUE in non-growing seasons; and (vi) combined temperature and precipitation changes had greater impacts on GPP/ET/WUE than individual changes. HIGHLIGHTS The SRPCM was proposed for parameter optimization of the BIOME-BGC model.; The variations of GPP, ET and WUE in 2003–2019 were explored on multiple time-scales.; GPP, ET and WUE in growing seasons played a decisive role in annual GPP and ET.; Daily WUE within 0–2 g/kg dominated in both non-/growing seasons during 2003–2019.; The combined rise of temperature and precipitation has greater impacts on GPP/ET/WUE than only temperature or precipitation rise.

Ghost: A General Framework for High-Performance Online Similarity Queries over Distributed Trajectory Streams

Trajectory similarity queries, including similarity search and similarity join, offer a foundation for many geo-spatial applications. With the rapid increase of streaming trajectory data volumes, e.g., data from mobile phones, vessel monitoring, or traffic systems, many location-based services benefit from online similarity analytics over trajectory data streams, where moving objects continually emit real-time position data. However, most existing studies focus on offline settings, and thus several major challenges remain unanswered in an online setting. To this end, we describe Ghost, a distributed stream processing framework that enables generic, efficient, and scalable online trajectory similarity search and join.We propose a novel incremental online similarity computation (IOSC) mechanism to accelerate pair-wise streaming trajectory distance calculation, which supports a broad range of trajectory distance metrics. Compared with previous studies, IOSC reduces the complexity from quadratic to linear in terms of trajectory length. Building on this foundation, we propose histogram-based algorithms that exploit histogram indexes and a series of pruning bounds to enable streaming trajectory similarity search and join. Finally, we extend our methods to the distributed platform Flink for scalability, where a CostPartitioner is developed to ensure parallel processing and workload balancing. An experimental study using two real-life and one synthetic datasets shows that Ghost (i) acquires 6-20× efficiency/throughput gains and one order of magnitude memory overhead savings over state-of-the-art baselines, (ii) achieves 3--8× workload balancing gains on Flink, and (iii) exhibits low parameter sensitivity and high robustness

Rheological Behavior and Microstructure Characteristics of SCC Incorporating Metakaolin and Silica Fume

This study explores the effects of metakaolin (MK) and silica fume (SF) on rheological behaviors and microstructure of self-compacting concrete (SCC). The rheology, slump flow, V-funnel, segregation degree (SA), and compressive strength of SCC are investigated. Microstructure characteristics, including hydration product and pore structure, are also studied. The results show that adding MK and SF instead of 4%, 6% and 8% fly ash (FA) reduces flowability of SCC; this is due to the fact that the specific surface area of MK and SF is larger than FA, and the total water demand increases as a result. However, the flowability increases when replacement ratio is 2%, as the small MK and SF particles will fill in the interstitial space of mixture and more free water is released. The fluidity, slump flow, and SA decrease linearly with the increase of yield stress. The total amount of SF and MK should be no more than 6% to meet the requirement of self-compacting. Adding MK or SF to SCC results in more hydration products, less Ca(OH)2 and refinement of pore structure, leading to obvious strength and durability improvements. When the total dosage of MK and SF admixture is 6%, these beneficial effects on workability, mechanical performance, and microstructure are more significant when SF and MK are applied together

Anti-Inflammatory Effects of Chloranthalactone B in LPS-Stimulated RAW264.7 Cells

Chloranthalactone B (CTB), a lindenane-type sesquiterpenoid, was obtained from the Chinese medicinal herb Sarcandra glabra, which is frequently used as a remedy for inflammatory diseases. However, the anti-inflammatory mechanisms of CTB have not been fully elucidated. In this study, we investigated the molecular mechanisms underlying these effects in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. CTB strongly inhibited the production of nitric oxide and pro-inflammatory mediators such as prostaglandin E2, tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β), and IL-6 in RAW264.7 cells stimulated with LPS. A reverse-transcription polymerase chain reaction assay and Western blot further confirmed that CTB inhibited the expression of inducible nitric oxide synthase, cyclooxygenase-2, TNF-α, and IL-1β at the transcriptional level, and decreased the luciferase activities of activator protein (AP)-1 reporter promoters. These data suggest that inhibition occurred at the transcriptional level. In addition, CTB blocked the activation of p38 mitogen-activated protein kinase (MAPK) but not c-Jun N-terminal kinase or extracellular signal-regulated kinase 1/2. Furthermore, CTB suppressed the phosphorylation of MKK3/6 by targeting the binding sites via formation of hydrogen bonds. Our findings clearly show that CTB inhibits the production of inflammatory mediators by inhibiting the AP-1 and p38 MAPK pathways. Therefore, CTB could potentially be used as an anti-inflammatory agent

Deqi

Deqi sensation, a psychophysical response characterized by a spectrum of different needling sensations, is essential for Chinese acupuncture clinical efficacy. Previous research works have investigated the component of Deqi response upon acupuncture on acupoints on the trunk and limbs. However, the characteristics of Deqi sensations of transcutaneous electrical nerve stimulation (TENS) on auricular points are seldom reported. In this study, we investigated the individual components of Deqi during TENS on auricular concha area and the superior scapha using quantitative measurements in the healthy subjects and depression patients. The most striking characteristics of Deqi sensations upon TENS on auricular points were tingling, numbness, and fullness. The frequencies of pressure, warmness, heaviness, and soreness were relatively lower. The dull pain and coolness are rare. The characteristics of Deqi were similar for the TENS on concha and on the superior scapha