Peroxynitrite is a damaging agent of oxidative stress that has been difficult to monitor in living cells. Here, an isatin-based chemiluminescent probe for peroxynitrite is reported.

Azanone (HNO) is a reactive nitrogen species with pronounced biological activity and high therapeutic potential for cardiovascular dysfunction. A critical barrier to understanding the biology of HNO and furthering clinical development is the quantification and real‐time monitoring of its delivery in living systems. Herein, we describe the design and synthesis of the first chemiluminescent probe for HNO, HNOCL‐1, which can detect HNO generated from concentrations of Angeli's salt as low as 138 nm with high selectivity based on the reaction with a phosphine group to form a self‐cleavable azaylide intermediate. We have capitalized on this high sensitivity to develop a generalizable kinetics‐based approach, which provides real‐time quantitative measurements of HNO concentration at the picomolar level. HNOCL‐1 can monitor dynamics of HNO delivery in living cells and tissues, demonstrating the versatility of this method for tracking HNO in living systems.

Iron is an essential nutrient for life, and its capacity to cycle between different oxidation states is required for processes spanning oxygen transport and respiration to nucleotide synthesis and epigenetic regulation. However, this same redox ability also makes iron, if not regulated properly, a potentially dangerous toxin that can trigger oxidative stress and damage. New methods that enable monitoring of iron in living biological systems, particularly in labile Fe forms, can help identify its contributions to physiology, aging, and disease. In this review, we summarize recent developments in activity-based sensing (ABS) probes for fluorescence Fe detection.

The metabolism of ethanol to acetaldehyde has been visualized in living lung epithelial cells using a hydrazinyl naphthalimide fluorescent probe. Utilizing a condensation reaction between carbonyls and a hydrazine moeity, we demonstrate that the fluorescent probe (Aldehydefluor-1) AF1 reacts with a range of reactive carbonyl species including formaldehyde, acetaldehyde, glyoxylic acid, and methyl glyoxal. With AF1, it is possible to directly visualize endogenous carbonyl metabolites. Here, we have applied it towards the visualization of acetaldehyde generated from alcohol dehydrogenase mediated ethanol metabolism, validating it as a useful tool to study the roles of alcohol in respiratory disease and other pathological mechanisms.

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Azanone (HNO) is ar eactive nitrogen species with pronounced biological activity and high therapeutic potential for cardiovascular dysfunction. Ac ritical barrier to understanding the biology of HNO and furthering clinical development is the quantification and real-time monitoring of its delivery in living systems.H erein, we describe the design and synthesis of the first chemiluminescent probe for HNO, HNOCL-1,w hich can detect HNO generated from concentrations of Angeliss alt as lowa s1 38 nm with high selectivity based on the reaction with ap hosphine group to form as elfcleavable azaylide intermediate.W eh ave capitalized on this high sensitivity to develop ag eneralizable kinetics-based approach,whichprovides real-time quantitative measurements of HNO concentration at the picomolar level. HNOCL-1 can monitor dynamics of HNO delivery in living cells and tissues, demonstrating the versatility of this method for tracking HNO in living systems.Azanone (HNO,n itroxyl), is chemically related to nitric oxide (NO) by the addition of one electron and one proton. HNO rapidly dimerizes and eliminates to form nitrous oxide (k = 8 10 6 m À1 s À1 at 23 8 8C) [1] and reacts with molecular oxygen with estimated rate constants that range from 10 3 to 10 4 m À1 s À1 (k = 3 10 3 m À1 s À1 at 37 8 8C; [2] k = 1 10 4 m À1 s À1 at 23 8 8C; [3] k = 1.8 10 4 m À1 s À1 at 25 8 8C [4,5] ). Tw omajor biological targets responsible for HNO bioactivity are thiols,which react directly with HNO to form sulfinamides (k = 2 10 6 m À1 s À1 at 37 8 8C) [2] and the iron in heme-containing proteins. [6] While both HNO and NO can activate soluble guanylyl cyclase,only HNO acts apositive cardiac inotrope by direct reaction with cysteine residues on cardiac ryanodine receptors and the sarcoplasmic reticulum Ca 2+

Activity-based protein profiling (ABPP) is a versatile strategy for identifying and characterizing functional protein sites and compounds for therapeutic development. However, the vast majority of ABPP methods for covalent drug discovery target highly nucleophilic amino acids such as cysteine or lysine. Here, we report a methionine-directed ABPP platform using Redox-Activated Chemical Tagging (ReACT), which leverages a biomimetic oxidative ligation strategy for selective methionine modification. Application of ReACT to oncoprotein cyclin-dependent kinase 4 (CDK4) as a representative high-value drug target identified three new ligandable methionine sites. We then synthesized a methionine-targeting covalent ligand library bearing a diverse array of heterocyclic, heteroatom, and stereochemically rich substituents. ABPP screening of this focused library identified 1oxF11 as a covalent modifier of CDK4 at an allosteric M169 site. This compound inhibited kinase activity in a dose-dependent manner on purified protein and in breast cancer cells. Further investigation of 1oxF11 found prominent cation-π and H-bonding interactions stabilizing the binding of this fragment at the M169 site. Quantitative mass-spectrometry studies validated 1oxF11 ligation of CDK4 in breast cancer cell lysates. Further biochemical analyses revealed cross-talk between M169 oxidation and T172 phosphorylation, where M169 oxidation prevented phosphorylation of the activating T172 site on CDK4 and blocked cell cycle progression. By identifying a new mechanism for allosteric methionine redox regulation on CDK4 and developing a unique modality for its therapeutic intervention, this work showcases a generalizable platform that provides a starting point for engaging in broader chemoproteomics and protein ligand discovery efforts to find and target previously undruggable methionine sites.

The undergraduate transfer process has well-documented challenges, especially for those who identify with groups historically excluded from science, technology, engineering, and mathematics (STEM) programs. Because transfer students gain later access to university networking and research opportunities than first-time-in-college students, transfer students interested in pursuing postbaccalaureate degrees in chemistry have a significantly shortened timeline in which to conduct research, a crucial component in graduate school applications. Mentorship programs have previously been instituted as effective platforms for the transfer of community cultural wealth within large institutions. We report here the design, institution, and assessment of a near-peer mentorship program for transfer students, the Transfer Student Mentorship Program (TSMP). Founded in 2020 by graduate students, the TSMP pairs incoming undergraduate transfer students with current graduate students for personalized mentorship and conducts discussion-based seminars to foster peer relationships. The transfer student participants have access to a fast-tracked networking method during their first transfer semester that can serve as a route for acquiring undergraduate research positions. Program efficacy was assessed via surveys investigating the rates of research participation and sense of belonging of transfer students. We observed that respondents that participated in the program experienced an overall improvement in these measures compared to respondents who did not. Having been entirely designed, instituted, and led by graduate students, we anticipate that this program will be highly tractable to other universities looking for actionable methods to improve their students’ persistence in pursuing STEM degrees.