An Activity‐Based Sensing Approach for the Detection of Cyclooxygenase‐2 in Live Cells

Yadav, Anuj; Reinhardt, Christopher J.; Arango, Andres S.; Huff, Hannah C.; Dong, Liang; Malkowski, Michael G.; Das, Aditi; Tajkhorshid, Emad; Chan, Jefferson

doi:10.1002/anie.201914845

Cited by 48 publications

(32 citation statements)

References 83 publications

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“…CoxFluor was validated as as ystem for inhibitor validation and for the detection of COX-2 activity within live cells using confocal microscopy and flow cytometry.F inally,C oxFluor was employed for the identification of oxygen-dependent activity changes that where not ar esult of protein expression ( Figure 4d). [47] This example highlights the importance of ABS for studying protein function and regulation beyond expression and translation. Cytochrome P450s (CYPs) catalyze the heme-dependent oxygenation of av ariety of metabolites and xenobiotics.…”

Section: Cyclooxygenasementioning

confidence: 97%

Advances in Activity‐Based Sensing Probes for Isoform‐Selective Imaging of Enzymatic Activity

2020

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Until recently, there were no generalizable methods for assessing the effects of post‐translational regulation on enzymatic activity. Activity‐based sensing (ABS) has emerged as a powerful approach for monitoring small‐molecule and enzyme activities within living systems. Initial examples of ABS were applied for measuring general enzymatic activity; however, a recent focus has been placed on increasing the selectivity to monitor a single enzyme or isoform. The highest degree of selectivity is required for differentiating between isoforms, where the targets display significant structural similarities as a result of a gene duplication or alternative splicing. This Minireview highlights key examples of small‐molecule isoform‐selective probes with a focus on the relevance of isoform differentiation, design strategies to achieve selectivity, and applications in basic biology or in the clinic.

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Section: Cyclooxygenasementioning

confidence: 97%

Advances in Activity‐Based Sensing Probes for Isoform‐Selective Imaging of Enzymatic Activity

2020

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“…31 We envision that activity-based fluorescent sensors can address this gap. [32][33][34][35][36][37] This strategy affords the ability to chemically tune and transform an enzyme's substrate into a fluorescent imaging platform for live-cell applications. To our knowledge, activity-based sensing has not been widely exploited for phenol sulfotransferases in living cells.…”

Section: Main Textmentioning

confidence: 99%

An Activity-Based Fluorescent Sensor for the Detection of the Phenol Sulfotransferase SULT1A1 in Living Cells

Baglia

Mills

Mitra

et al. 2020

Preprint

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<p>The biological activation and incorporation of inorganic sulfate proceeds via a process known as sulfurylation. Transfer of a sulfuryl moiety from the activated sulfate donor, 3’-phosphoadenosine-5’-phosphosulfate (PAPS), to hydroxy-containing substrates by human phenol sulfotransferases (SULT1 family) alters substrate solubility and charge to affect the metabolism of endogenous metabolites, xenobiotics, and drugs. Current methods to monitor SULT1 activity in living cells primarily rely on radiolabeling and/or cell extractions, but these methods do not provide a direct readout of enzyme activity with a dynamic, temporally resolved spatial map in live, intact cells. To fill this gap, here, we present the development, computational modeling, <i>in vitro</i> enzymology, and biological application of Sulfotransferase Sensor-3, STS-3, an activity-based fluorescent sensor for SULT1A1, the most widely expressed and promiscuous SULT1 isoform. </p>

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“…Recently,Z hang [43,44] and Hong [45] et al embarked on the survey of the physical environment inside aggregated proteins,b ut their work primarily revealed the changes in polarity and viscosity upon protein aggregation using environment-sensitive probes.H owever,i ts till remains elusive how chemically reactive proteins are at their aggregated state. Therefore,c urrent powerful chemical sensing and bioconjugation strategies [46][47][48] may provide hints to address this question.…”

Section: Introductionmentioning

confidence: 99%

Covalent Probes for Aggregated Protein Imaging via Michael Addition

et al. 2021

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Covalent chemical reactions to modify aggregated proteins are rare.Here,wereported covalent Michael addition can generally occur upon protein aggregation. Suchr eactivity was initially discovered by ab ioinspired fluorescent colorswitch probe mimicking the photo-conversion mechanism of Kaede fluorescent protein. This probe was dark with folded proteins but turned on red fluorescence (620 nm) when it noncovalently bound to misfolded proteins.S upported by the biochemical and mass spectrometry results,t he probe chemoselectively reacted with the reactive cysteines of aggregated proteins via covalent Michael addition and gradually switched to green fluorescence (515 nm) upon protein aggregation. Exploiting this Michael addition chemistry in the malachite green dye derivatives demonstrated its general applicability and chemicalt unability,r esulting in different fluorescence color-switch responses.O ur work may offer an ew avenue to explore other chemical reactions upon protein aggregation and design covalent probes for imaging,c hemical proteomics,and therapeutic purposes.

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An Activity‐Based Sensing Approach for the Detection of Cyclooxygenase‐2 in Live Cells

Cited by 48 publications

References 83 publications

Advances in Activity‐Based Sensing Probes for Isoform‐Selective Imaging of Enzymatic Activity

Advances in Activity‐Based Sensing Probes for Isoform‐Selective Imaging of Enzymatic Activity

An Activity-Based Fluorescent Sensor for the Detection of the Phenol Sulfotransferase SULT1A1 in Living Cells

Covalent Probes for Aggregated Protein Imaging via Michael Addition

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