<i>S</i>‐Nitrosothiol Chemistry at the Single‐Molecule Level

Choi, Lai-Sheung; Bayley, Hagan

doi:10.1002/ange.201202365

Cited by 12 publications

(9 citation statements)

References 40 publications

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“…By contrast, the model reaction of GSNO with GSH in vitro supposes both N-and S-directed thiolate attack to rationalize myriad observed products; however, the control exerted by protein R-groups on SNO-generating thiol reactions is absent in this model, and low-yield S-directed reactions are observed because of high reactant concentrations (167,200). Reaction of GSNO with an engineered protein nanopore primarily resulted in transnitrosylation reactions at physiological pH (36). In addition, SNAP does not participate in S-thiolation reactions, likely due to R-group steric hindrance (36).…”

Section: Protein Sno-thiol Reaction Biasmentioning

confidence: 90%

Protein S-Nitrosylation: Determinants of Specificity and Enzymatic Regulation of S-Nitrosothiol-Based Signaling

Stomberski

Hess

Stamler

2019

Antioxidants & Redox Signaling

208

180

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The emerging picture of NO biology entails equilibria among potentially thousands of different SNOs, governed by denitrosylases and nitrosylases. Thus, to elucidate the operation and consequences of S-nitrosylation in cellular contexts, studies should consider the roles of SNO-proteins as both targets and transducers of S-nitrosylation, functioning according to enzymatically governed equilibria. Antioxid. Redox Signal. 00, 000-000.

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Section: Protein Sno-thiol Reaction Biasmentioning

confidence: 90%

Protein S-Nitrosylation: Determinants of Specificity and Enzymatic Regulation of S-Nitrosothiol-Based Signaling

Stomberski

Hess

Stamler

2019

Antioxidants & Redox Signaling

208

180

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“…1). The P C band was cut out and rehydrated, and the protein was extracted (18). Purified proteins were stored at -80 C in TE buffer (10 mM Tris.HCl, 1 mM EDTA, pH 8.0) containing 5 mM dithiothreitol (DTT).…”

Section: Protein Preparationmentioning

confidence: 99%

Rates and Stoichiometries of Metal Ion Probes of Cysteine Residues within Ion Channels

Choi

Mach

Bayley

2013

Biophysical Journal

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Metal ion probes are used to assess the accessibility of cysteine side chains in polypeptides lining the conductive pathways of ion channels and thereby determine the conformations of channel states. Despite the widespread use of this approach, the chemistry of metal ion-thiol interactions has not been fully elucidated. Here, we investigate the modification of cysteine residues within a protein pore by the commonly used Ag(+) and Cd(2+) probes at the single-molecule level, and provide rates and stoichiometries that will be useful for the design and interpretation of accessibility experiments.

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“…Single-molecule chemistry has attracted interest because it provides insights into fundamental chemical processes that cannot be observed at the ensemble scale (6)(7)(8). However, chemistry carried out within the αHL nanoreactor has so far been confined to the reactions of thiolates (i.e., deprotonated Cys side chains) (7)(8)(9)(10)(11)(12) and derivatives of the side chains of Cys residues (13,14). The incorporation of unnatural amino acids into the αHL pore would substantially advance single-molecule chemistry, through the incorporation of a large variety of individual side chains as well as multiple substitutions.…”

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confidence: 99%

Semisynthetic protein nanoreactor for single-molecule chemistry

Lee

Bayley

2015

Proc. Natl. Acad. Sci. U.S.A.

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The covalent chemistry of individual reactants bound within a protein pore can be monitored by observing the ionic current flow through the pore, which acts as a nanoreactor responding to bond-making and bond-breaking events. In the present work, we incorporated an unnatural amino acid into the α-hemolysin (αHL) pore by using solid-phase peptide synthesis to make the central segment of the polypeptide chain, which forms the transmembrane β-barrel of the assembled heptamer. The full-length αHL monomer was obtained by native chemical ligation of the central synthetic peptide to flanking recombinant polypeptides. αHL pores with one semisynthetic subunit were then used as nanoreactors for single-molecule chemistry. By introducing an amino acid with a terminal alkyne group, we were able to visualize click chemistry at the single-molecule level, which revealed a long-lived (4.5-s) reaction intermediate. Additional side chains might be introduced in a similar fashion, thereby greatly expanding the range of single-molecule covalent chemistry that can be investigated by the nanoreactor approach.semisynthesis | alpha-hemolysin | single-molecule chemistry | click chemistry

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S‐Nitrosothiol Chemistry at the Single‐Molecule Level

Cited by 12 publications

References 40 publications

Protein S-Nitrosylation: Determinants of Specificity and Enzymatic Regulation of S-Nitrosothiol-Based Signaling

Protein S-Nitrosylation: Determinants of Specificity and Enzymatic Regulation of S-Nitrosothiol-Based Signaling

Rates and Stoichiometries of Metal Ion Probes of Cysteine Residues within Ion Channels

Semisynthetic protein nanoreactor for single-molecule chemistry

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