The electroweak corrections at one-loop level to the process H\ensuremath{\rightarrow}tt\ifmmode\bar\else\textasciimacron\fi{} are calculated, incorporating the top mass value announced recently by CDF. For ${\mathit{M}}_{\mathit{H}}$1 TeV where a perturbative calculation is valid, the corrections themselves would gain a few to 20% increment in the decay width as the Higgs-boson mass ${\mathit{M}}_{\mathit{H}}$ is increasing within the region, but they are in opposite direction to the QCD ones. If the electroweak and QCD corrections are considered together, the resultant decay width of the mode yields a reduction about a few percent of the tree level one.
Experimental explorations of functional mechanisms in natural electron-transfer proteins are often frustrated by their fragility and extreme complexity. We have designed and synthesized four-α-helix-bundle redox proteins, maquettes, that are much simplified and more robust than natural redox proteins and can be designed to bind onto electrode surfaces to facilitate systematic investigations. The points of interest that can be now assessed are not only the processes that govern biological assembly of equilibrium structures, electrochemistry, and electron tunneling rates but also how these factors are coupled together to effect redox driven catalysis. Here we describe maquettes that bis-histidine ligate protoporphyrin IX (heme), much like native b cytochromes, as well as contain charged surface patches, much like native cytochrome c. The positively charged residues aid adsorption to negatively charged surfaces, such as gold electrodes modified by 11-mercaptoundecanoic acid, and facilitate cyclic voltammetry (CV) measurements. CV demonstrates the reversible electrochemistry typical for cytochrome b as well as the coupling of the b-heme oxidation and reduction to proton exchange. The pH dependency of redox midpoint potentials reveals a major (three pH units) shift of the pKa which matches the shift previously shown to originate in nearby glutamates1. The redox potentials correspondingly shift from −0.24 (pH > pKred, deprotonated) to −0.11 V (pH < pKox, protonated). The rate of electron transfer at zero driving force between the hemes and the gold electrode was determined to be 120 s-1, a rate consistent with tunneling through the mercaptoundecanoic acid spacer and suggesting that the coupled proton exchange is not rate limiting. Reduction of the heme in the presence of CO-saturated buffer shifted the oxidation peak from −0.2 to +0.35 V, indicating massive preferential CO binding to the reduced heme. Consistent with solution spectroscopy, CO must displace one axial histidine to the heme to form the His−CO form of the ferrous heme. The CO is released upon heme oxidation at high potentials. In contrast to coupled proton exchange, CO binding/release and ligand exchange are slow on the time scale of electron tunneling between the heme edge and the electrode.
We apply X-ray interferometry to study the profile structure of Langmuir−Blodgett (LB) monolayers containing maquette peptides, de novo di-α-helical synthetic peptides designed as model systems for studying biological electron transfer. The results demonstrate that it is possible to create monolayers with the peptide vectorially oriented with its helical axis (the direction of electron transfer within the holopeptide) approximately normal to the surface of the solid support. This orientation can even be achieved when the orientation of the peptide in the precursor Langmuir film at the air/water interface is parallel to the surface, indicating that reorganization of the monolayer can occur during or after LB deposition. Though issues regarding the low density of the film and variability between samples remain to be addressed, the work represents an important step toward future correlated functional/structural studies of these peptides.
We sought to construct a monolayer composed of four-α-helical bundles formed by association of di-α-helical peptides. Specular X-ray reflectivity showed that dihelices that have been made amphiphilic by attachment of C16 hydrocarbon chains to their N-termini can be vectorially oriented in Langmuir monolayers at an air−water interface with their helical axes normal to the interface. But off-specular X-ray reflectivity indicated that these dihelices did not associate to form four-helix bundles possibly because they were constrained to be of parallel topology. To achieve four-helix bundles vectorially oriented at the interface, we relaxed this constraint to allow for a 1:1 association of the amphiphilic dihelices with their water-soluble counterparts. Electron density profiles for the monolayers derived from specular X-ray reflectivity demonstrated four-helix bundle formation only when the association between dihelices is directed via designed attractive electrostatic interactions between the polar faces of the amphipathic helices.
This paper describes the use of surface plasmon resonance spectroscopy and self-assembled monolayers (SAMs) of alkanethiols on gold to evaluate the ability of surfaces terminating in different combinations of charged groups to resist the nonspecific adsorption of proteins from aqueous buffer. Mixed SAMs formed from a 1:1 combination of a thiol terminated in a trimethylammonium group and a thiol terminated in a sulfonate group adsorbed less than 1% of a monolayer of two proteins with different characteristics: fibrinogen and lysozyme. Single-component SAMs formed from thiols terminating in groups combining a positively charged moiety and a negatively charged moiety were also capable of resisting the adsorption of proteins. Single-component SAMs presenting single charges adsorbed nearly a full monolayer of protein. The amount of protein that adsorbed to mixed zwitterionic SAMs did not depend on the ionic strength or the pH of the buffer in which the protein was dissolved. The amount of protein that adsorbed to single-component zwitterionic SAMs increased as the ionic strength of the buffer decreased; it also decreased as the pH of the buffer increased (at constant ionic strength). Single-component zwitterionic SAMs composed of thiols terminating in N,N-dimethyl-amino-propane-1-sulfonic acid (−N+(CH3)2CH2CH2CH2SO3-) groups were substantially more effective at resisting adsorption of fibrinogen and lysozyme from buffer at physiological ionic strength and pH than single-component zwitterionic SAMs composed of thiols terminating in phosphoric acid 2-trimethylamino-ethyl ester (−OP(O)2-OCH2CH2N+(CH3)3). Several of these zwitterionic SAMs were comparable to the best known systems for resisting nonspecific adsorption of protein.
An electrical junction formed by mechanical contact between two self-assembled monolayers (SAMs)a SAM formed from an dialkyl disulfide with a covalently linked tetracyanoquinodimethane group that is supported by silver (or gold) and a SAM formed from an alkanethiolate SAM that is supported by mercuryrectifies current. The precursor to the SAM on silver (or gold) was bis(20-(2-((2,5-cyclohexadiene-1,4-diylidene)dimalonitrile))decyl)) disulfide and that for the SAM on mercury was HS(CH2)n-1CH3 (n = 14, 16, 18). The electrical properties of the junctions were characterized by current−voltage measurements. The ratio of the conductivity of the junction in the forward bias (Hg cathodic) to that in the reverse bias (Hg anodic), at a potential of 1 V, was 9 ± 2 when the SAM on mercury was derived from HS(CH2)15CH3. The ratio of the conductivity in the forward bias to that in the reverse bias increased with decreasing chain length of the alkanethiol used to form the SAM on mercury. These results demonstrate that a single redox center asymmetrically placed in a metal−insulator−metal junction can cause the rectification of current and indicate that a fixed dipole in the insulating region of a metal−insulator−metal junction is not required for rectification.
A new method for improving low-concentration sample recovery and reducing sample preparation steps in matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) is presented. In the conventional approach, samples are typically desalted and/or concentrated with various techniques and deposited on the MALDI target as small droplets. In this work, we describe a new approach in which an elastomeric device is reversibly sealed on the MALDI target to form a multi-well plate with the MALDI target as the base of the plate. The new format allows a larger volume (5-200 microL) of samples to be deposited on each spot and a series of sample handling processes, including desalting and concentrating, to be performed directly on the MALDI target. Several advantages have been observed: (i) multiple sample transferring steps are avoided; (ii) recovery of low-concentration peptides during sample preparation is improved using a novel desalting method that utilizes the hydrophobic surface of the elastomeric device; and (iii) sequence coverage of the peptide mass fingerprinting map is improved using a novel method in which proteins are immobilized on the hydrophobic surface of the elastomeric device for in-well trypsin digestion, followed by desalting and concentrating the digestion products in the same well.
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