Two-dimensional covalent organic frameworks (2D-COFs) are a class of crystalline porous organic polymers that consist of covalently linked, two-dimensional sheets that can stack together through noncovalent interactions. Here we report the synthesis of a novel COF, called PyCOFamide, which has an experimentally observed pore size that is greater than 6 nm in diameter. This is among the largest pore size reported to date for a 2D-COF. PyCOFamide exhibits permanent porosity and high crystallinity as evidenced by the nitrogen adsorption, powder X-ray diffraction, and high-resolution transmission electron microscopy. We show that the pore size of PyCOFamide is large enough to accommodate fluorescent proteins such as Superfolder green fluorescent protein and mNeonGreen. This work demonstrates the utility of noncovalent structural reinforcement in 2D-COFs to produce larger and persistent pore sizes than previously possible.

Natural and laboratory-guided evolution has created a rich diversity of fluorescent protein (FP)-based sensors for chloride (Cl−). To date, such sensors have been limited to the Aequorea victoria green fluorescent...

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Nuclear magnetic resonance (NMR) spectroscopy is vital to synthesis and provides rich problem-solving opportunities to organic chemistry students. Using the theories of scaffolding, interleaving, and blocking, our research systematically explores how textbooks introduce and reinforce spectral features when teaching students to solve ¹H NMR spectroscopy problems. Specifically, we investigated the ¹H NMR spectral features presented in worked examples and practice problems across four undergraduate organic chemistry textbooks. We examined the frequency and ordering of spectral features to explore how the textbooks could support scaffolded instruction. Spectral features like the number of signals and chemical shift were covered by problems more frequently, while integration was covered least. Our findings suggest that textbooks do not provide sufficient practice with all ¹H NMR spectral features. We observed no discernible pattern in how textbooks ordered spectral features of ¹H NMR in problems, indicating that there is little systematic method to the design of textbook chapters. Implications for textbook authors and editors, instruction, and research are discussed.

Using the principles of supramolecular design, we demonstrate that a urea-functionalized porous organic polymer can recognize anionic dyes in water.

Chloride is a ubiquitous ion essential for regulating biological processes. Long thought to be just a counterion in biological systems, chloride is emerging as a vital anion that plays a major role in a range of diseases. The ability to visualize intracellular chloride has been enabled by fluorescent protein-based sensors. Although the yellow fluorescent protein from Aequorea victoria, and variants thereof, quench in the presence of chloride and other anions, they have provided valuable insights in biological studies and have established the benchmark for applying protein-based sensors for chloride. However, a single domain fluorescent protein sensor that generates a turn-on response to provide a spatial and temporal map of endogenous chloride has yet to be reported. Here, we highlight how mutagenesis of non-coordinating residues in the binding pocket of mNeonGreen has unlocked this new sensing capability. Our protein engineering efforts coupled with in vitro spectroscopy have led us to discover the ChlorON series as turn-on fluorescent sensors for chloride that operate at physiological pH. Fluorescence microscopy experiments with the ChlorON sensors reveal that cells have baseline levels of chloride, which could not have been directly observed with turn-off sensors. These advancements set the stage for visualizing endogenous chloride dynamics in real-time and uncovering new roles for chloride in biology.

Detection of anions in complex aqueous media is a fundamental challenge with practical utility that can be addressed by supramolecular chemistry. Biomolecular hosts such as proteins can be used and adapted as an alternative to synthetic hosts. Here, we report how the mutagenesis of the β-bulge residues (D137 and W138) in mNeonGreen, a bright, monomeric fluorescent protein, unlocks and tunes the anion preference at physiological pH for sulfate, resulting in the turn-off sensor SulfOFF-1. This unprecedented sensing arises from an enhancement in the kinetics of binding, largely driven by position 138. In line with these data, molecular dynamics (MD) simulations capture how the coordinated entry and gating of sulfate into the β-barrel is eliminated upon mutagenesis to facilitate binding and fluorescence quenching.

Detection of anions in complex aqueous media is a fundamental challenge with practical utility that can be addressed by supramolecular chemistry. Biomolecular hosts such as proteins can be used and adapted as an alternative to synthetic hosts. Here, we report how the mutagenesis of the β‐bulge residues (D137 and W138) in mNeonGreen, a bright, monomeric fluorescent protein, unlocks and tunes the anion preference at physiological pH for sulfate, resulting in the turn‐off sensor SulfOFF‐1. This unprecedented sensing arises from an enhancement in the kinetics of binding, largely driven by position 138. In line with these data, molecular dynamics (MD) simulations capture how the coordinated entry and gating of sulfate into the β‐barrel is eliminated upon mutagenesis to facilitate binding and fluorescence quenching.