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1. Substrate-Dependent Sensitivity of SIRT1 to Nicotinamide Inhibition

   https://doi.org/10.3390/biom11020312  

   Rymarchyk, Stacia; Kang, Wenjia; Cen, Yana (February 2021, Biomolecules)

   null (Ed.)

   SIRT1 is the most extensively studied human sirtuin with a broad spectrum of endogenous targets. It has been implicated in the regulation of a myriad of cellular events, such as gene transcription, mitochondria biogenesis, insulin secretion as well as glucose and lipid metabolism. From a mechanistic perspective, nicotinamide (NAM), a byproduct of a sirtuin-catalyzed reaction, reverses a reaction intermediate to regenerate NAD+ through “base exchange”, leading to the inhibition of the forward deacetylation. NAM has been suggested as a universal sirtuin negative regulator. Sirtuins have evolved different strategies in response to NAM regulation. Here, we report the detailed kinetic analysis of SIRT1-catalyzed reactions using endogenous substrate-based synthetic peptides. A novel substrate-dependent sensitivity of SIRT1 to NAM inhibition was observed. Additionally, SIRT1 demonstrated pH-dependent deacetylation with normal solvent isotope effects (SIEs), consistent with proton transfer in the rate-limiting step. Base exchange, in contrast, was insensitive to pH changes with no apparent SIEs, indicative of lack of proton transfer in the rate-limiting step. Consequently, NAM inhibition was attenuated at a high pH in proteated buffers. Our study provides new evidence for “activation by de-repression” as an effective sirtuin activation strategy. 

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2. Oxime as a general photocage for the design of visible light photo-activatable fluorophores

   https://doi.org/10.1039/d1sc05351e  

   Wang, Lushun; Wang, Shichao; Tang, Juan; Espinoza, Vanessa B.; Loredo, Axel; Tian, Zeru; Weisman, R. Bruce; Xiao, Han (December 2021, Chemical Science)

   Photoactivatable fluorophores have been widely used for tracking molecular and cellular dynamics with subdiffraction resolution. In this work, we have prepared a series of photoactivatable probes using the oxime moiety as a new class of photolabile caging group in which the photoactivation process is mediated by a highly efficient photodeoximation reaction. Incorporation of the oxime caging group into fluorophores results in loss of fluorescence. Upon light irradiation in the presence of air, the oxime-caged fluorophores are oxidized to their carbonyl derivatives, restoring strong fluorophore fluorescence. To demonstrate the utility of these oxime-caged fluorophores, we have created probes that target different organelles for live-cell confocal imaging. We also carried out photoactivated localization microscopy (PALM) imaging under physiological conditions using low-power light activation in the absence of cytotoxic additives. Our studies show that oximes represent a new class of visible-light photocages that can be widely used for cellular imaging, sensing, and photo-controlled molecular release. 

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3. C–H bond activation via concerted metalation–deprotonation at a palladium( iii ) center

   https://doi.org/10.1039/D3SC00034F  

   Bouley, Bailey S.; Tang, Fengzhi; Bae, Dae Young; Mirica, Liviu M. (April 2023, Chemical Science)

   Herein we report the direct observation of C–H bond activation at an isolated mononuclear Pd( iii ) center. The oxidation of the Pd( ii ) complex ( Me N4)Pd II (neophyl)Cl (neophyl = –CH 2 C(CH 3 ) 2 Ph; Me N4 = N , N ′-dimethyl-2,11-diaza[3.3](2,6)pyridinophane) using the mild oxidant ferrocenium hexafluorophosphate (FcPF 6 ) yields the stable Pd( iii ) complex [( Me N4)Pd III (neophyl)Cl]PF 6 . Upon the addition of an acetate source, [( Me N4)Pd III (neophyl)Cl]PF 6 undergoes Csp 2 –H bond activation to yield the cyclometalated product [( Me N4)Pd III (cycloneophyl)]PF 6 . This metalacycle can be independently prepared, allowing for a complete characterization of both the starting and final Pd( iii ) complexes. The C–H activation step can be monitored directly by EPR and UV-Vis spectroscopies, and kinetic isotope effect (KIE) studies suggest that either a pre-association step such as an agostic interaction may be rate limiting, or that the C–H activation is partially rate-limiting in conjunction with ligand rearrangement. Density functional theory calculations support that the reaction proceeds through a κ 3 ligand coordination and that the flexible ligand structure is important for this transformation. Overall, this study represents the first example of discrete C–H bond activation occurring at a Pd( iii ) center through a concerted metalation–deprotonation mechanism, akin to that observed for Pd( ii ) and Pd( iv ) centers. 

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4. Integrating Machine Learning and Color Chemistry: Developing a High-School Curriculum toward Real-World Problem-Solving

   https://doi.org/10.1021/acs.jchemed.3c00589  

   Jiang, Shiyan; McClure, Jeanne; Mao, Hongjing; Chen, Jiahui; Liu, Yunshu; Zhang, Yang (December 2023, Journal of Chemical Education)

   Artificial intelligence (AI) is rapidly transforming our world, making it imperative to educate the next generation about both the potential benefits and the challenges associated with AI. This study presents a cross-disciplinary curriculum that connects AI and chemistry disciplines in the high school classroom. Particularly, we leverage machine learning (ML), an important and simple application of AI to instruct students to build an ML-based virtual pH meter for high-precision pH read-outs. We used a “codeless” and free ML neural network building software, Orange, along with a simple chemical topic of pH to show the connection between AI and chemistry for high-schoolers who might have rudimentary backgrounds in both disciplines. The goal of this curriculum is to promote student interest and drive in the analytical chemistry domain and offer insights into how the interconnection between chemistry and ML can benefit high-school students in science learning. The activity involves students using pH strips to measure the pH of various solutions with local relevancy and then building an ML neural network model to predict the pH value based on the color changes of pH strips. The integrated curriculum increased student interest in chemistry and ML and demonstrated the relevance of science to students’ daily lives and global issues. This approach is transformative in developing a broad spectrum of integration topics between chemistry and ML and understanding their global impacts. 

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5. Droplet Microfluidic Technology for the Early and Label-Free Isolation of Highly-Glycolytic, Activated T-Cells

   https://doi.org/10.3390/mi13091442  

   Zielke, Claudia; Gutierrez Ramirez, Adriana J.; Voss, Kelsey; Ryan, Maya S.; Gholizadeh, Azam; Rathmell, Jeffrey C.; Abbyad, Paul (September 2022, Micromachines)

   A label-free, fixation-free and passive sorting method is presented to isolate activated T-cells shortly after activation and prior to the display of activation surface markers. It uses a recently developed sorting platform dubbed “Sorting by Interfacial Tension” (SIFT) that sorts droplets based on pH. After polyclonal (anti-CD3/CD28 bead) activation and a brief incubation on chip, droplets containing activated T-cells display a lower pH than those containing naive cells due to increased glycolysis. Under specific surfactant conditions, a change in pH can lead to a concurrent increase in droplet interfacial tension. The isolation of activated T-cells on chip is hence achieved as flattened droplets are displaced as they encounter a micro-fabricated trench oriented diagonally with respect to the direction of flow. This technique leads to an enrichment of activated primary CD4+ T-cells to over 95% from an initial mixed population of naive cells and cells activated for as little as 15 min. Moreover, since the pH change is correlated to successful activation, the technique allows the isolation of T-cells with the earliest activation and highest glycolysis, an important feature for the testing of T-cell activation modulators and to determine regulators and predictors of differentiation outcomes. 

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