Peptide Tag/Probe Pairs Based on the Coordination Chemistry for Protein Labeling

Uchinomiya, Shohei; Ojida, Akio; Hamachi, Itaru

doi:10.1021/ic401612z

Cited by 32 publications

(25 citation statements)

References 42 publications

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“…Ala-scanning mutational analysis revealed significant effects (SI Figure S9) only from combined mutagenesis of D100, D102 that comprise the DxD and T139 (found in its accessory ion-engagement site 25 , entries 10,11,13). Although other backbone interactions that cannot be 'scanned' in this way likely also contribute, the partial role of this combined site was confirmed by triple mutation (entries 14,15) 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 tive binding/basic residues, e.g. H217, R131, had essentially no effect (Table 1).…”

Section: Scheme 1 Established Strategies Compared To Metalsite Guidementioning

confidence: 96%

Selective Metal-Site-Guided Arylation of Proteins

et al. 2016

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We describe palladium-mediated S-arylation that exploits natural metal-binding motifs to ensure high site selectivity for a proximal reactive residue. This allows the chemical identification not only of proteins that bind metals but also the environment of the metal-binding site itself through proteomic analysis of arylation sites. The transformation is easy to perform under standard conditions, does not require the isolation of a reactive Ar-Pd complex, is broad in scope, and is applicable in cell lysates as well as to covalent inhibition/modulation of metal-dependent enzymatic activity.

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Section: Scheme 1 Established Strategies Compared To Metalsite Guidementioning

confidence: 96%

Selective Metal-Site-Guided Arylation of Proteins

et al. 2016

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“…Various NTA-based fluorescent probes have been developed via conjugation of fluorophores with mono-NTA (12,17) or di-, tri-, and tetra-NTA derivatives to either mimic the concept of FlAsH or overcome the weak binding nature of His-tag with Ni 2+ -NTA (K d = 13 μM) (11,(18)(19)(20)(21). Despite significant increases in stability of the multiple chelator heads/His-tag adducts compared with mono-NTA, negative charges of these moieties might prevent them from entering cells.…”

Section: +mentioning

confidence: 99%

Rapid labeling of intracellular His-tagged proteins in living cells

Lai¹,

Chang²,

Hu³

et al. 2015

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

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Small molecule-based fluorescent probes have been used for real-time visualization of live cells and tracking of various cellular events with minimal perturbation on the cells being investigated. Given the wide utility of the (histidine)6-Ni2+-nitrilotriacetate (Ni-NTA) system in protein purification, there is significant interest in fluorescent Ni2+-NTA–based probes. Unfortunately, previous Ni-NTA–based probes suffer from poor membrane permeability and cannot label intracellular proteins. Here, we report the design and synthesis of, to our knowledge, the first membrane-permeable fluorescent probe Ni-NTA-AC via conjugation of NTA with fluorophore and arylazide followed by coordination with Ni2+ ions. The probe, driven by Ni2+-NTA, binds specifically to His-tags genetically fused to proteins and subsequently forms a covalent bond upon photoactivation of the arylazide, leading to a 13-fold fluorescence enhancement. The arylazide is indispensable not only for fluorescence enhancement, but also for strengthening the binding between the probe and proteins. Significantly, the Ni-NTA-AC probe can rapidly enter different types of cells, even plant tissues, to target His-tagged proteins. Using this probe, we visualized the subcellular localization of a DNA repair protein, Xeroderma pigmentosum group A (XPA122), which is known to be mainly enriched in the nucleus. We also demonstrated that the probe can image a genetically engineered His-tagged protein in plant tissues. This study thus offers a new opportunity for in situ visualization of large libraries of His-tagged proteins in various prokaryotic and eukaryotic cells.

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“…More refined protein tags for the selective labeling of intracellular proteins have recently been developed based on advances in synthetic chemistry (Jing and Cornish, 2011;Uchinomiya et al, 2014). For example, SNAP-tag is a peptide of 182 amino acids that is derived from the human O 6 -alkylguanine-DNA alkyltransferase.…”

Section: Cali With Transgenically Encoded Tagsmentioning

confidence: 99%

Chromophore-assisted laser inactivation – towards a spatiotemporal–functional analysis of proteins, and the ablation of chromatin, organelle and cell function

Sano

Watanabe

Matsunaga

2014

Journal of Cell Science

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Chromophore-assisted laser or light inactivation (CALI) has been employed as a promising technique to achieve spatiotemporal knockdown or loss-of-function of target molecules in situ. CALI is performed using photosensitizers as generators of reactive oxygen species (ROS). There are two CALI approaches that use either transgenic tags with chemical photosensitizers, or genetically encoded fluorescent protein fusions. Using spatially restricted microscopy illumination, CALI can address questions regarding, for example, protein isoforms, subcellular localization or phase-specific analyses of multifunctional proteins that other knockdown approaches, such as RNA interference or treatment with chemicals, cannot. Furthermore, rescue experiments can clarify the phenotypic capabilities of CALI after the depletion of endogenous targets. CALI can also provide information about individual events that are involved in the function of a target protein and highlight them in multifactorial events. Beyond functional analysis of proteins, CALI of nuclear proteins can be performed to induce cell cycle arrest, chromatin-or locus-specific DNA damage. Even at organelle levelsuch as in mitochondria, the plasma membrane or lysosomes -CALI can trigger cell death. Moreover, CALI has emerged as an optogenetic tool to switch off signaling pathways, including the optical depletion of individual neurons. In this Commentary, we review recent applications of CALI and discuss the utility and effective use of CALI to address open questions in cell biology.

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Peptide Tag/Probe Pairs Based on the Coordination Chemistry for Protein Labeling

Cited by 32 publications

References 42 publications

Selective Metal-Site-Guided Arylation of Proteins

Selective Metal-Site-Guided Arylation of Proteins

Rapid labeling of intracellular His-tagged proteins in living cells

Chromophore-assisted laser inactivation – towards a spatiotemporal–functional analysis of proteins, and the ablation of chromatin, organelle and cell function

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