Selective Fluorescence Detection of Small‐Molecule‐Binding Proteins by Using a Dual Photoaffinity Labeling System

Sakurai, Kaori; Yamada, Ryuji; Okada, Ayumi; Tawa, Masaki; Ozawa, Shutaro; Inoue, Maia

doi:10.1002/cbic.201200758

Cited by 13 publications

(13 citation statements)

References 66 publications

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“…A similar approach was devised to differentiate specific and nonspecific labeling by Sakurai et al, 147 developing a dual PAL system. If the active and inactive photoprobes are structurally similar, both structures are assumed to bind nonspecifically to the same proteins or domains (Figure 58).…”

Section: Chemical Reviewsmentioning

confidence: 99%

The Life of Pi Star: Exploring the Exciting and Forbidden Worlds of the Benzophenone Photophore

et al. 2016

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The widespread applications of benzophenone (BP) photochemistry in biological chemistry, bioorganic chemistry, and material science have been prominent in both academic and industrial research. BP photophores have unique photochemical properties: upon n-π* excitation at 365 nm, a biradicaloid triplet state is formed reversibly, which can abstract a hydrogen atom from accessible C-H bonds; the radicals subsequently recombine, creating a stable covalent C-C bond. This light-directed covalent attachment process is exploited in many different ways: (i) binding/contact site mapping of ligand (or protein)-protein interactions; (ii) identification of molecular targets and interactome mapping; (iii) proteome profiling; (iv) bioconjugation and site-directed modification of biopolymers; (v) surface grafting and immobilization. BP photochemistry also has many practical advantages, including low reactivity toward water, stability in ambient light, and the convenient excitation at 365 nm. In addition, several BP-containing building blocks and reagents are commercially available. In this review, we explore the "forbidden" (transitions) and excitation-activated world of photoinduced covalent attachment of BP photophores by touring a colorful palette of recent examples. In this exploration, we will see the pros and cons of using BP photophores, and we hope that both novice and expert photolabelers will enjoy and be inspired by the breadth and depth of possibilities.

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Section: Chemical Reviewsmentioning

confidence: 99%

The Life of Pi Star: Exploring the Exciting and Forbidden Worlds of the Benzophenone Photophore

et al. 2016

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“…7e). The authors have performed a proof-of-principle study with this active/ inactive dual-probe system (Sakurai et al 2013).…”

Section: Active/inactive Dual-probe Systemmentioning

confidence: 99%

“…Typically, eluted samples are resolved on one dimension gel. In order to improve resolution, researchers have also utilized 2-dimension electrophoresis (Brehmer et al 2004;Jessen et al 2005;Sakurai et al 2013). However, accurately identifying specific protein bands from gel images is still a difficult task.…”

Section: Two-dimensional Gel Electrophoresis (Fitge)mentioning

confidence: 99%

Affinity purification in target identification: the specificity challenge

Zheng

2015

Arch. Pharm. Res.

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Since phenotype-based screening directly evaluates capability of small molecules for modulating biology in actual biological systems, it has become an important discover modality in modern pharmaceutical sciences. However, in order to fully elucidate the molecular mechanism underlying the bioactivity of small molecules, identification of their biological targets is an indispensable step. Among the many target identification strategies developed during the past several decades, affinity purification remains to be one of the most important and powerful approaches, as it can directly reveal the physical interactions between small molecules and their biomolecular targets. However, due to the complexity of the proteome and the diversity of small molecule-protein interactions, affinity purification faces the specificity challenge: how to identify the true specific targets from the non-specific background? Focusing on this challenge, in this review, we briefly introduce the history and background of affinity purification, and then we discussed the major technological developments aiming to address this challenge. We have summarized these approaches in two categories: noise reduction and comparative distinction. This review also highlights the importance of choosing an integrated approach combining multiple methods to achieving success in target identification.

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“…Compounds 2 and 4 represented inactive analogs. A similar approach was used by the same group [58], using an active/inactive dual-PAL system for selective crosslinking and straightforward detection of small-molecule-binding proteins. They designed a novel PAL reaction in which nonspecific proteins were scavenged by an inactive probe and co-reacted with a conventional PAL probe.…”

Section: Benzophenone-based Probes To Study Ppimentioning

confidence: 99%

Photo-affinity labeling (PAL) in chemical proteomics: a handy tool to investigate protein-protein interactions (PPIs)

et al. 2016

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Protein-protein interactions (PPIs) trigger a wide range of biological signaling pathways that are crucial for biomedical research and drug discovery. Various techniques have been used to study specific proteins, including affinity chromatography, activity-based probes, affinity-based probes and photo-affinity labeling (PAL). PAL has become one of the most powerful strategies to study PPIs. Traditional photocrosslinkers are used in PAL, including benzophenone, aryl azide, and diazirine. Upon photoirradiation, these photocrosslinkers (Pls) generate highly reactive species that react with adjacent molecules, resulting in a direct covalent modification. This review introduces recent examples of chemical proteomics study using PAL for PPIs.

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Selective Fluorescence Detection of Small‐Molecule‐Binding Proteins by Using a Dual Photoaffinity Labeling System

Cited by 13 publications

References 66 publications

The Life of Pi Star: Exploring the Exciting and Forbidden Worlds of the Benzophenone Photophore

The Life of Pi Star: Exploring the Exciting and Forbidden Worlds of the Benzophenone Photophore

Affinity purification in target identification: the specificity challenge

Photo-affinity labeling (PAL) in chemical proteomics: a handy tool to investigate protein-protein interactions (PPIs)

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