- Rapid acidolysis of benzyl group as a suitable approach for syntheses of peptides naturally produced by oxidative stress and containing 3-nitrotyrosine
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3-Nitrotyrosine (Nit) belongs to products of oxidative stress and could probably influence conformation of neurodegenerative proteins. Syntheses of peptides require availability of suitable synthon for introduction of Nit residue. Common phenolic protecti
- Niederhafner, Petr,?afarík, Martin,Brichtová, Eva,?ebestík, Jaroslav
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p. 1087 - 1098
(2016/04/05)
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- Kinetics of 3-nitrotyrosine modification on exposure to hypochlorous acid
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The markers 3-nitrotyrosine and 3-chlorotyrosine are measured as surrogates for reactive nitrogen species and hypochlorous acid respectively, which are both elevated in inflamed human tissues. Previous studies reported a loss of 3-nitrotyrosine when exposed to hypochlorous acid, suggesting that observations of 3-nitrotyrosine underestimate the presence of reactive nitrogen species in diseased tissue (Whiteman and Halliwell, Biochemical and Biophysical Research Communications, 258, 168-172 (1999)). This report evaluates the significance of 3-nitrotyrosine loss by measuring the kinetics of the reaction between 3-nitrotyrosine and hypochlorous acid. The results demonstrate that 3-nitrotyrosine is chlorinated by hypochlorous acid or chloramines to form 3-chloro-5-nitrotyrosine. As 3-nitrotyrosine from in vivo samples is usually found within proteins rather than as free amino acid, we also examined the reaction of 3-nitrotyrosine modification in the context of peptides. The chlorination of 3-nitrotyrosine in peptides was observed to occur up to 700-fold faster than control reactions using equivalent amino acid mixtures. These results further advance our understanding of tyrosine chlorination and the use of 3-nitrotyrosine formed in vivo as a biomarker of reactive nitrogen species.
- Curtis,Neidigh
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p. 1355 - 1362
(2015/01/08)
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- Real-time monitoring of the dephosphorylating activity of protein tyrosine phosphatases using microarrays with 3-nitrophosphotyrosine substrates
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Phosphatases and kinases regulate the crucial phosphorylation post-translational modification. In spite of their similarly important role in many diseases and therapeutic potential, phosphatases have received arguably less attention. One reason for this is a scarcity of high-throughput phosphatase assays. Herein, a new real-time, dynamic protein tyrosine phosphatase (PTP) substrate microarray assay measuring product formation is described. PTP substrates comprising a novel 3-nitrophosphotyrosine residue are immobilized in discrete spots. After reaction catalyzed by a PTP a 3-nitrotyrosine residue is formed that can be detected by specific, sequence-independent antibodies. The resulting microarray was successfully evaluated with a panel of recombinant PTPs and cell lysates, which afforded results comparable to data from other assays. Its parallel nature, convenience, and low sample requirements facilitate investigation of the therapeutically relevant PTP enzyme family. Keeping it real: The activity of important protein tyrosine phosphatases has been monitored in real time in parallel with a novel substrate microarray through formation of 3-nitrotyrosine (see figure). Copyright
- Van Ameijde, Jeroen,Overvoorde, John,Knapp, Stefan,Den Hertog, Jeroen,Ruijtenbeek, Rob,Liskamp, Rob M. J.
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p. 1349 - 1357
(2013/12/04)
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- Discovery of a novel nonphosphorylated pentapeptide motif displaying high affinity for Grb2-SH2 domain by the utilization of 3′-substituted tyrosine derivatives
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The growth factor receptor-bound protein 2 (Grb2) is an SH2 domain-containing docking module that represents an attractive target for anticancer therapeutic intervention. An impressive number of synthetic Grb2-SH2 domain inhibitors have been identified; however, clinical agents operating by this mechanism are lacking, due in part to the unique requirement of anionic phosphate-mimicking functionality for high SH2 domain-binding affinity or the extended peptide nature of most inhibitors. In the current study, a new binding motif was successfully developed by the incorporation of 3′-substituted tyrosine derivatives into a simplified nonphosphorylated cyclic pentapeptide scaffold (4), which resulted in high affinity Grb2-SH2 inhibitors without any phosphotyrosine or phosphotyrosine mimetics. The new L-amino acid analogues bearing an additional nitro, amino, hydroxy, methoxy or carboxy group at the 3′-position of the phenol ring of tyrosine were prepared in an orthogonally protected form suitable for solid-phase peptide synthesis using Fmoc protocols. The incorporation of these residues into cyclic peptides composed of a five-amino acid sequence motif, Xx′-Leu-(3′- substituted-Tyr)-Ac6c-Asn, provided a brand new class of nonphosphorylated Grb2 SH2 domain inhibitors with reduced size, charge and peptidic character. The highest binding affinity was exhibited by the 3′-aminotyrosine (3′-NH2-Tyr)-containing (R)-sulfoxide-cyclized pentapeptide (10b) with an IC50 = 58 nM, the first example with low-nanomolar affinity for a five-amino acid long sequence binding to Grb2-SH2 domain free of any phosphotyrosine or phosphotyrosine mimics. However, the incorporation of 3′-NO2-Tyr, 3′-OH-Tyr or 3′-OCH3-Tyr surrogates in the pentapeptide scaffold is detrimental to Grb2-SH2 binding. These observations were rationalized using molecular modeling. More significantly, the best Grb2-SH2 inhibitor 10b showed excellent activity in inhibiting the growth of erbB2-dependent MDA-MB-453 tumor cell lines with an IC50 value of 19 nM. This study is the first attempt to identify novel nonphosphorylated high affinity Grb2 SH2 inhibitors by the utilization of 3′-substituted tyrosine derivatives, providing a promising new strategy and template for the development of non-pTyr-containing Grb2-SH2 domain antagonists with potent cellular activity, which potentially may find value in chemical therapeutics for erbB2-related cancers.
- Song, Yan-Li,Peach, Megan L.,Roller, Peter P.,Qiu, Su,Wang, Shaomeng,Long, Ya-Qiu
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p. 1585 - 1596
(2007/10/03)
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- Development of L-3-aminotyrosine suitably protected for the synthesis of a novel nonphosphorylated hexapeptide with low-nanomolar Grb2-SH2 domain-binding affinity
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Synthesis of orthogonally protected (2S)-2-amino-3-(3-amino-4-hydroxy- phenyl)-propionic acid (10) suitable for solid phase peptide synthesis and its first use for the preparation of nonphosphorylated Grb2-SH2 domain antagonists (4a-c) are reported. The 3-aminotyrosine containing sulfoxide-cyclized hexapeptide (4b) exhibited potent Grb2-SH2 domain binding affinity with IC 50=50nM, which represents the highest affinity yet reported for a peptide inhibitor against Grb2-SH2 domain with only 6 residues free of phosphotyrosine or phosphotyrosine mimics. This potent small peptidomimetic 4b may be representative of a new class of therapeutically relevant Grb2-SH2 domain-directed agents, and acts as a chemotherapeutic lead for the treatment of erbB2-related cancers.
- Song, Yan-Li,Roller, Peter P.,Long, Ya-Qiu
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p. 3205 - 3208
(2007/10/03)
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