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11.
Application of plug–plug technique to ACE experiments for discovery of peptides binding to a larger target protein: A model study of calmodulin‐binding fragments selected from a digested mixture of reduced BSA
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Kazuki Saito Mamiko Nakato Takaaki Mizuguchi Shinji Wada Hiromasa Uchimura Hiroshi Kataoka Shigeyuki Yokoyama Hiroshi Hirota Yoshiaki Kiso 《Electrophoresis》2014,35(6):846-854
To discover peptide ligands that bind to a target protein with a higher molecular mass, a concise screening methodology has been established, by applying a “plug–plug” technique to ACE experiments. Exploratory experiments using three mixed peptides, mastoparan‐X, β‐endorphin, and oxytocin, as candidates for calmodulin‐binding ligands, revealed that the technique not only reduces the consumption of the protein sample, but also increases the flexibility of the experimental conditions, by allowing the use of MS detection in the ACE experiments. With the plug–plug technique, the ACE–MS screening methodology successfully selected calmodulin‐binding peptides from a random library with diverse constituents, such as protease digests of BSA. Three peptides with Kd values between 8–147 μM for calmodulin were obtained from a Glu‐C endoprotease digest of reduced BSA, although the digest showed more than 70 peaks in its ACE–MS electropherogram. The method established here will be quite useful for the screening of peptide ligands, which have only low affinities due to their flexible chain structures but could potentially provide primary information for designing inhibitors against the target protein. 相似文献
12.
Shigeyuki Namiki Kenji Takikawa Mako Kamiya Prof. Tetsuo Nagano Prof. Yasuteru Urano Prof. Kenzo Hirose 《Angewandte Chemie (International ed. in English)》2014,53(24):6085-6089
Live imaging of exocytosis dynamics is crucial for a precise spatiotemporal understanding of secretion phenomena, but current approaches have serious limitations. We designed and synthesized small‐molecular fluorescent probes that were chemically optimized for sensing acidic intravesicular pH values, and established that they can be used to sensitively and reliably visualize vesicular dynamics following stimulation. This straightforward technique for the visualization of exocytosis as well as endocytosis/reacidification processes with high spatiotemporal precision is expected to be a powerful tool for investigating dynamic cellular phenomena involving changes in the pH value. 相似文献
13.
Dr. Masaki Yoshida Dr. Mio Kondo Prof. Toshikazu Nakamura Prof. Ken Sakai Prof. Shigeyuki Masaoka 《Angewandte Chemie (International ed. in English)》2014,53(43):11519-11523
A series of [{(terpy)(bpy)Ru}(μ‐O){Ru(bpy)(terpy)}]n+ ( [RuORu]n+ , terpy=2,2′;6′,2′′‐terpyridine, bpy=2,2′‐bipyridine) was systematically synthesized and characterized in three distinct redox states (n=3, 4, and 5 for RuII,III2 , RuIII,III2 , and RuIII,IV2 , respectively). The crystal structures of [RuORu]n+ (n=3, 4, 5) in all three redox states were successfully determined. X‐ray crystallography showed that the Ru? O distances and the Ru‐O‐Ru angles are mainly regulated by the oxidation states of the ruthenium centers. X‐ray crystallography and ESR spectra clearly revealed the detailed electronic structures of two mixed‐valence complexes, [RuIIIORuIV]5+ and [RuIIORuIII]3+ , in which each unpaired electron is completely delocalized across the oxo‐bridged dinuclear core. These findings allow us to understand the systematic changes in structure and electronic state that accompany the changes in the redox state. 相似文献
14.
Tozuka Z Kaneko H Shiraga T Mitani Y Beppu M Terashita S Kawamura A Kagayama A 《Journal of mass spectrometry : JMS》2003,38(8):793-808
Triple-stage quadrupole (TSQ) electrospray ionization (ESI) tandem mass spectrometry (MS/MS) and ion trap ESI-MS/MS can be used to cleave protonated molecules to produce carbocations and neutral molecules in the positive ion mode. Dissociation products which correspond to protonated forms of neutral fragment molecules can also be trapped and detected. These protonated molecules in turn can cleave via carbocation cleavage, ipso cleavage, onium cleavage or McLafferty or related rearrangements. One can elucidate the structures of metabolites from the differences in m/z ratios of the fragments arising from the original drug compound and its metabolite. This strategy for structural elucidation is further facilitated by estimates of the reactivity of drugs with oxygen diradicals involved in cytochrome P-450 cycles. 相似文献
15.
Shigeyuki Mayama Toshikazu Tani Tamio Ueno Kazumasa Hirabayashi Tadakazu Nakashima Hiroshi Fukami Yukio Mizuno Hiroshi Irie 《Tetrahedron letters》1981,22(22):2103-2106
The major phytoalexin from oat leaves has been identified as 2-[2-(4-hydroxyphenyDethenyl] -6-hydroxy-4H-3,1-benzoxazin-4-one (). 相似文献
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Philippe De Taxis Du Poet Shigeyuki Miyamoto Toru Murakami Jun Kimura Isao Karube 《Analytica chimica acta》1990
Direct electron transfer between active glucose oxidase (GOD) and a gold electrode was obtained when GOD was immobilized in poly(N-methylpyrrole) electrochemically prepared on the gold electrode. When electropolymerization was accomplished at 50 °C, after glucose addition, the cyclic voltammograms showed an increased oxidation peak at ca. ?0.45 V vs. Ag/AgCl. This potential corresponds to the oxidation potential for FADH2. Although the GOD becomes much less selective, a glucose-dependent current response is obtained. 相似文献
19.
Kazunari Ozasa Shigeyuki Nemoto Mizuo Maeda Masahiko Hara 《Physica E: Low-dimensional Systems and Nanostructures》2006,32(1-2):89
We have found that the photoluminescence (PL) intensity of CdSe/ZnS nanocrystals placed on a thin film of insulator (GaAsOx/GaAs) depends on excitation wavelength through the interference effects of the excitation light. By employing the multi-reflection/interference calculation, the insulator thickness of the underlying non-uniform patterns can be evaluated by the simple observation of CdSe/ZnS PL with a couple of excitation wavelengths. Moreover, the differences observed for the temporal evolution of CdSe/ZnS PL (blue shifts and degradation) among the excitation wavelengths suggest that the photo-induced changes of chemical composition and surface ligands are responsible for blue shifts and degradation, respectively. 相似文献
20.
Yamauchi K Masaoka S Sakai K 《Dalton transactions (Cambridge, England : 2003)》2011,40(46):12447-12449
To answer the question of whether Pt(II)-based H(2)-evolving catalysts are stable upon exposure to H(2), the behaviours of some platinum(II) complexes in the presence of H(2) (5 × 10(-4) - 1 atm) have been followed spectrophotometrically. The results reveal that some catalysts are highly stable upon exposure to H(2). 相似文献