Catalytic Alcoholysis of Tetramethylsilane via Pt-Mediated C−H Bond Activation

Tetramethylsilane reacts with 2,2,2-trifluoroethanol (TFE) in the presence of a cationic platinum(II) catalyst [(NN)PtMe(TFE)]+ (NN = 1,2-bis(3,5-dimethylphenylimino)butane). Catalytic Si−C bond heterolysis results in the formation of the trimethylsilyl ether, Me_3SiOCH_2CF_3, accompanied by liberation of one equivalent of methane. Preliminary experiments suggest that a rate-determining C−H bond activation precedes rapid attack by solvent at silicon to yield the silyl ether product and regenerate the active platinum methyl cation

The role of alkane coordination in C–H bond cleavage at a Pt(II) center

The rates of CFormula H bond activation for various alkanes by [(N–N)Pt(Me)(TFEd3)]+ (N Formula N = ArFormula NFormula C(Me)Formula C(Me)Formula NFormula Ar; Ar = 3,5-di-tert-butylphenyl; TFE-d3 = CF3CD2OD) were studied. Both linear and cyclic alkanes give the corresponding alkene-hydride cation [(N–N)Pt(H)(alkene)]+ via (i) rate determining alkane coordination to form a CFormula H {sigma} complex, (ii) oxidative cleavage of the coordinated CFormula H bond to give a platinum(IV) alkyl-methyl-hydride intermediate, (iii) reductive coupling to generate a methane {sigma} complex, (iv) dissociation of methane, and (v) beta-H elimination to form the observed product. Second-order rate constants for cycloalkane activation (CnH2n), are proportional to the size of the ring (k ~ n). For cyclohexane, the deuterium kinetic isotope effect (kH/kD) of 1.28 (5) is consistent with the proposed rate determining alkane coordination to form a CFormula H {sigma} complex. Statistical scrambling of the five hydrogens of the Pt-methyl and the coordinated methylene unit, via rapid, reversible steps ii and iii, and interchange of geminal CFormula H bonds of the methane and cyclohexane CFormula H {sigma} adducts, is observed before loss of methane

Kinetic and Thermodynamic Preferences in Aryl vs Benzylic C−H Bond Activation with Cationic Pt(II) Complexes

Anhydrous cationic Pt(II) complexes [(NN)Pt(CH_3)(CF_3CD_2OD)]+ (1, NN = ArNC(Me)−C(Me)NAr), which are obtained by reaction of (NN)Pt(CH_3)_2 with B(C_6F_5)_3 in CF_3CD_2OD, activate C−H bonds of benzene and methylbenzenes, with enhanced reactivity compared to the previously prepared equilibrium mixtures with the (thermodynamically favored) aquo complexes. For methylbenzenes (toluene, p-xylene, mesitylene), activation at the aromatic and benzylic positions are kinetically competitive, but the product of the latter is strongly favored thermodynamically. This unusual trend is attributed to formation of η^3-benzyl structures, which can be observed spectroscopically for 1,4-diethylbenzene activation

Studying the Salt Dependence of the Binding of σ70 and σ32 to Core RNA Polymerase Using Luminescence Resonance Energy Transfer

The study of protein-protein interactions is becoming increasingly important for understanding the regulation of many cellular processes. The ability to quantify the strength with which two binding partners interact is desirable but the accurate determination of equilibrium binding constants is a difficult process. The use of Luminescence Resonance Energy Transfer (LRET) provides a homogeneous binding assay that can be used for the detection of protein-protein interactions. Previously, we developed an LRET assay to screen for small molecule inhibitors of the interaction of σ70 with theβ' coiled-coil fragment (amino acids 100–309). Here we describe an LRET binding assay used to monitor the interaction of E. coli σ70 and σ32 with core RNA polymerase along with the controls to verify the system. This approach generates fluorescently labeled proteins through the random labeling of lysine residues which enables the use of the LRET assay for proteins for which the creation of single cysteine mutants is not feasible. With the LRET binding assay, we are able to show that the interaction of σ70 with core RNAP is much more sensitive to NaCl than to potassium glutamate (KGlu), whereas the σ32 interaction with core RNAP is insensitive to both salts even at concentrations >500 mM. We also find that the interaction of σ32 with core RNAP is stronger than σ70 with core RNAP, under all conditions tested. This work establishes a consistent set of conditions for the comparison of the binding affinities of the E.coli sigma factors with core RNA polymerase. The examination of the importance of salt conditions in the binding of these proteins could have implications in both in vitro assay conditions and in vivo function

X-ray absorption spectroscopy systematics at the tungsten L-edge

A series of mononuclear six-coordinate tungsten compounds spanning formal oxidation states from 0 to +VI, largely in a ligand environment of inert chloride and/or phosphine, has been interrogated by tungsten L-edge X-ray absorption spectroscopy. The L-edge spectra of this compound set, comprised of [W<sup>0</sup>(PMe<sub>3</sub>)<sub>6</sub>], [W<sup>II</sup>Cl<sub>2</sub>(PMePh<sub>2</sub>)<sub>4</sub>], [W<sup>III</sup>Cl<sub>2</sub>(dppe)<sub>2</sub>][PF<sub>6</sub>] (dppe = 1,2-bis(diphenylphosphino)ethane), [W<sup>IV</sup>Cl<sub>4</sub>(PMePh<sub>2</sub>)<sub>2</sub>], [W<sup>V</sup>(NPh)Cl<sub>3</sub>(PMe<sub>3</sub>)<sub>2</sub>], and [W<sup>VI</sup>Cl<sub>6</sub>] correlate with formal oxidation state and have usefulness as references for the interpretation of the L-edge spectra of tungsten compounds with redox-active ligands and ambiguous electronic structure descriptions. The utility of these spectra arises from the combined correlation of the estimated branching ratio (EBR) of the L<sub>3,2</sub>-edges and the L<sub>1</sub> rising-edge energy with metal Z<sub>eff</sub>, thereby permitting an assessment of effective metal oxidation state. An application of these reference spectra is illustrated by their use as backdrop for the L-edge X-ray absorption spectra of [W<sup>IV</sup>(mdt)<sub>2</sub>(CO)<sub>2</sub>] and [W<sup>IV</sup>(mdt)<sub>2</sub>(CN)<sub>2</sub>]<sup>2–</sup> (mdt<sup>2–</sup> = 1,2-dimethylethene-1,2-dithiolate), which shows that both compounds are effectively W<sup>IV</sup> species. Use of metal L-edge XAS to assess a compound of uncertain formulation requires: 1) Placement of that data within the context of spectra offered by unambiguous calibrant compounds, preferably with the same coordination number and similar metal ligand distances. Such spectra assist in defining upper and/or lower limits for metal Z<sub>eff</sub> in the species of interest; 2) Evaluation of that data in conjunction with information from other physical methods, especially ligand K-edge XAS; 3) Increased care in interpretation if strong π-acceptor ligands, particularly CO, or π-donor ligands are present. The electron-withdrawing/donating nature of these ligand types, combined with relatively short metal-ligand distances, exaggerate the difference between formal oxidation state and metal Z<sub>eff</sub> or, as in the case of [W<sup>IV</sup>(mdt)<sub>2</sub>(CO)<sub>2</sub>], add other subtlety by modulating the redox level of other ligands in the coordination sphere

The Cyclic AMP Cascade Is Altered in the Fragile X Nervous System

Fragile X syndrome (FX), the most common heritable cause of mental retardation and autism, is a developmental disorder characterized by physical, cognitive, and behavioral deficits. FX results from a trinucleotide expansion mutation in the fmr1 gene that reduces levels of fragile X mental retardation protein (FMRP). Although research efforts have focused on FMRP's impact on mGluR signaling, how the loss of FMRP leads to the individual symptoms of FX is not known. Previous studies on human FX blood cells revealed alterations in the cyclic adenosine 3′, 5′-monophosphate (cAMP) cascade. We tested the hypothesis that cAMP signaling is altered in the FX nervous system using three different model systems. Induced levels of cAMP in platelets and in brains of fmr1 knockout mice are substantially reduced. Cyclic AMP induction is also significantly reduced in human FX neural cells. Furthermore, cAMP production is decreased in the heads of FX Drosophila and this defect can be rescued by reintroduction of the dfmr gene. Our results indicate that a robust defect in cAMP production in FX is conserved across species and suggest that cAMP metabolism may serve as a useful biomarker in the human disease population. Reduced cAMP induction has implications for the underlying causes of FX and autism spectrum disorders. Pharmacological agents known to modulate the cAMP cascade may be therapeutic in FX patients and can be tested in these models, thus supplementing current efforts centered on mGluR signaling

A microfluidic-FCS platform for investigation on the dissociation of Sp1-DNA complex by doxorubicin

The transcription factor (TF) Sp1 is a well-known RNA polymerase II transcription activator that binds to GC-rich recognition sites in a number of essential cellular and viral promoters. In addition, direct interference of Sp1 binding to DNA cognate sites using DNA-interacting compounds may provide promising therapies for suppression of cancer progression and viral replication. In this study, we present a rapid, sensitive and cost-effective evaluation of a GC intercalative drug, doxorubicin (DOX), in dissociating the Sp1–DNA complex using fluorescence correlation spectroscopy (FCS) in a microfluidic system. FCS allows assay miniaturization without compromising sensitivity, making it an ideal analytical method for integration of binding assays into high-throughput, microfluidic platforms. A polydimethylsiloxane (PDMS)-based microfluidic chip with a mixing network is used to achieve specific drug concentrations for drug titration experiments. Using FCS measurements, the IC(50) of DOX on the dissociation of Sp1–DNA complex is estimated to be 0.55 μM, which is comparable to that measured by the electrophoretic mobility shift assay (EMSA). However, completion of one drug titration experiment on the proposed microfluidic-FCS platform is accomplished using only picograms of protein and DNA samples and less than 1 h total assay time, demonstrating vast improvements over traditional ensemble techniques

The Mere Exposure Effect in the Domain of Haptics

Background: Zajonc showed that the attitude towards stimuli that one had been previously exposed to is more positive than towards novel stimuli. This mere exposure effect (MEE) has been tested extensively using various visual stimuli. Research on the MEE is sparse, however, for other sensory modalities. Methodology/Principal Findings: We used objects of two material categories (stone and wood) and two complexity levels (simple and complex) to test the influence of exposure frequency (F0 = novel stimuli, F2 = stimuli exposed twice, F10 = stimuli exposed ten times) under two sensory modalities (haptics only and haptics & vision). Effects of exposure frequency were found for high complex stimuli with significantly increasing liking from F0 to F2 and F10, but only for the stone category. Analysis of ‘‘Need for Touch’ ’ data showed the MEE in participants with high need for touch, which suggests different sensitivity or saturation levels of MEE. Conclusions/Significance: This different sensitivity or saturation levels might also reflect the effects of expertise on the haptic evaluation of objects. It seems that haptic and cross-modal MEEs are influenced by factors similar to those in the visual domain indicating a common cognitive basis