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Gerhard Schenk Rosely A. Peralta Suzana Cimara Batista Adailton J. Bortoluzzi Bruno Szpoganicz Andrew K. Dick Paul Herrald Graeme R. Hanson Robert K. Szilagyi Mark J. Riley Lawrence R. Gahan Ademir Neves 《Journal of biological inorganic chemistry》2008,13(1):139-155
Purple acid phosphatases (PAPs) are a group of heterovalent binuclear metalloenzymes that catalyze the hydrolysis of phosphomonoesters
at acidic to neutral pH. While the metal ions are essential for catalysis, their precise roles are not fully understood. Here,
the Fe(III)Ni(II) derivative of pig PAP (uteroferrin) was generated and its properties were compared with those of the native
Fe(III)Fe(II) enzyme. The k
cat of the Fe(III)Ni(II) derivative (approximately 60 s−1) is approximately 20% of that of native uteroferrin, and the Ni(II) uptake is considerably faster than the reconstitution
of full enzymatic activity, suggesting a slow conformational change is required to attain optimal reactivity. An analysis
of the pH dependence of the catalytic properties of Fe(III)Ni(II) uteroferrin indicates that the μ-hydroxide is the likely
nucleophile. Thus, the Ni(II) derivative employs a mechanism similar to that proposed for the Ga(III)Zn(II) derivative of
uteroferrin, but different from that of the native enzyme, which uses a terminal Fe(III)-bound nucleophile to initiate catalysis.
Binuclear Fe(III)Ni(II) biomimetics with coordination environments similar to the coordination environment of uteroferrin
were generated to provide both experimental benchmarks (structural and spectroscopic) and further insight into the catalytic
mechanism of hydrolysis. The data are consistent with a reaction mechanism employing an Fe(III)-bound terminal hydroxide as
a nucleophile, similar to that proposed for native uteroferrin and various related isostructural biomimetics. Thus, only in
the uteroferrin-catalyzed reaction are the precise details of the catalytic mechanism sensitive to the metal ion composition,
illustrating the significance of the dynamic ligand environment in the protein active site for the optimization of the catalytic
efficiency.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
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Franck?CarboneroEmail authorView authors OrcID profile Alba?C.?Mayta-Apaza Jiang-Zhou?Yu Matt?Lindeblad Alex?Lyubimov Flavia?Neri Erzsebet?Szilagyi Amelia?Bartholomew 《Radiation and environmental biophysics》2018,57(4):419-426
In rodent studies, the gut microbiota has been implicated in facilitating both radioresistance, by protecting the epithelium from apoptotic responses and radiosensitivity, inducing endothelial apoptotic responses. Despite the observation that large animal models, such as the Chinese Rhesus macaque and the Gottingen Minipig, demonstrate similarity to human physiologic responses to radiation, little is known about radiation-induced changes of the gut microbiome in these models. To compare the two models, we used bioequivalent radiation doses which resulted in an LD50 for Gottingen Minipigs and Chinese Rhesus macaques, 1.9 Gy and 6.8 Gy, respectively. Fecal samples taken prior and 3 days post-radiation were used for 16S rRNA gene sequence amplicon high throughput sequencing (Illumina MiSeq). Baseline gut microbiota profiles were dissimilar between minipigs and rhesus macaques. Irradiation profoundly impacted gut microbiota profiles in both animals. Significant increases of intracellular symbionts were common to both models and to reported changes in rodents suggesting universality of these findings post-radiation. Remarkably, opposite dynamics were observed for the main phyla, with increase of Firmicutes and decrease of Bacteroidetes and Proteobacteria in minipigs but with enrichment of Bacteroidetes in rhesus macaques. Minipig changes in magnitude and in variety of species affected were more extensive than those observed in rhesus macaques. This pilot study provides an important first step in comparing the radiosensitive pig model to the comparatively more radioresistant macaque model, for the identification of microbial elements which may influence radiosensitivity. 相似文献
35.
Sodium depletion in dogs is known to affect both the renin-angiotensin as well as the sympathetic nervous system. The effect of this dietary regime upon the area postrema pressor pathway, as evaluated by the cardiovascular responses to centrally acting angiotensin II, has not been determined previously. With this in mind, male mongrel dogs were maintained on either a normal or a sodium restricted diet supplemented with furosemide and dose-response curves for intravertebral and intravenous angiotensin II (range: 1-20 ng/kg/min) were obtained. Sodium depletion results in not only a blunted intravenous pressor response to angiotensin II but also the abolition of the centrally mediated pressor responses mediated by the area postrema. Because accumulating evidence indicates that in sodium depleted dogs sympathetic nerve activity is reduced while central noradrenergic inhibitory activity is increased the reduced effects of angiotensin II upon the central sympathetically mediated pressor response may in part be related to decreases in sympathetic nerve activity. 相似文献
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Sen S Krishnakumar A McClead J Johnson MK Seefeldt LC Szilagyi RK Peters JW 《Journal of inorganic biochemistry》2006,100(5-6):1041-1052
In the present work, determination of the structure of the nitrogenase Leu 127 deletion variant Fe protein with MgATP bound is presented, along with density functional theory calculations, to provide insights into the roles of MgATP in the nitrogenase reaction mechanism. Comparison of the MgATP-bound structure of this Fe protein to the nucleotide-free form indicates that the binding of MgATP does not alter the overall structure of the variant significantly with only small differences in the conformation of amino acids in direct contact with the two bound MgATP molecules being seen. The earlier observation of splitting of the [4Fe-4S] cluster into two [2Fe-2S] clusters was observed to be unaltered upon binding MgATP. Density functional theory was used to probe the assignment of ligands to the two [2Fe-2S] rhombs. The Mg(2+) environment in the MgATP-bound structure of the Leu127 deletion Fe protein is similar to that observed for the Fe protein in the nitrogenase Fe protein: MoFe protein complex stabilized by MgADP and tetrafluoroaluminate suggesting that large scale conformational change implicated for the Fe protein may not be mediated by changes in the Mg(2+) coordination. The results presented here indicated that MgATP may enhance the stability of an open conformation and prohibit intersubunit interactions, which have been implicated in promoting nucleotide hydrolysis. This could be critical to the tight control of MgATP hydrolysis observed within the nitrogenase complex and may be important for maintaining unidirectional electron flow toward substrate reduction. 相似文献
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Kis-Toth K Hajdu P Bacskai I Szilagyi O Papp F Szanto A Posta E Gogolak P Panyi G Rajnavolgyi E 《Journal of immunology (Baltimore, Md. : 1950)》2011,187(3):1273-1280
Expression of CD1a protein defines a human dendritic cell (DC) subset with unique functional activities. We aimed to study the expression of the Nav1.7 sodium channel and the functional consequences of its activity in CD1a(-) and CD1a(+) DC. Single-cell electrophysiology (patch-clamp) and quantitative PCR experiments performed on sorted CD1a(-) and CD1a(+) immature DC (IDC) showed that the frequency of cells expressing Na(+) current, current density, and the relative expression of the SCN9A gene encoding Nav1.7 were significantly higher in CD1a(+) cells than in their CD1a(-) counterparts. The activity of Nav1.7 results in a depolarized resting membrane potential (-8.7 ± 1.5 mV) in CD1a(+) IDC as compared with CD1a(-) cells lacking Nav1.7 (-47 ± 6.2 mV). Stimulation of DC by inflammatory signals or by increased intracellular Ca(2+) levels resulted in reduced Nav1.7 expression. Silencing of the SCN9A gene shifted the membrane potential to a hyperpolarizing direction in CD1a(+) IDC, resulting in decreased cell migration, whereas pharmacological inhibition of Nav1.7 by tetrodotoxin sensitized the cells for activation signals. Fine-tuning of IDC functions by a voltage-gated sodium channel emerges as a new regulatory mechanism modulating the migration and cytokine responses of these DC subsets. 相似文献
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Galambas Amanda Miller Jacquelyn Jones Morgan McDaniel Elizabeth Lukes Molly Watts Hope Copié Valérie Broderick Joan B. Szilagyi Robert K. Shepard Eric M. 《Journal of biological inorganic chemistry》2019,24(6):793-807
JBIC Journal of Biological Inorganic Chemistry - The synthesis and characterization of short peptide-based maquettes of metalloprotein active sites facilitate an inquiry into their... 相似文献
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Travis V. Harris Robert K. Szilagyi Karen L. McFarlane Holman 《Journal of biological inorganic chemistry》2009,14(6):891-898
Several RuIII compounds are propitious anticancer agents although the precise mechanisms of action remain unknown. With this paper we start
to establish an experimental library of X-ray absorption spectroscopy (XAS) data for ten Ru compounds wherein the ligands
[Cl−, dimethyl sulfoxide, imidazole, and indazole] were varied systematically to provide electronic structural information for
future use in correlating spectroscopic signatures with chemical properties. Despite the considerable difference in the coordination
environments of the complexes studied, the overall differences in spectral features and electronic structures calculated using
density functional theory are unexpectedly small. However, the differences in the electronic structure of the RuIII prodrugs KP1019 ([IndH][trans-RuCl4(Ind)2], Ind is indazole) and ICR ([ImH][trans-RuCl4(Im)2], Im is imidazole) observed in the XAS data show correlation with known chemical and biological activities in addition to
the donor abilities of imidazole compared with indazole and reduction potentials of the complexes. These semiquantitative
results lay the groundwork for future biochemical studies into the structure–function relationships of Ru-based anticancer
drugs.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献