A ternary mechanism for NADH oxidation by positively charged electron acceptors, catalyzed at the flavin site in respiratory complex I

AbstractThe flavin mononucleotide in complex I (NADH:ubiquinone oxidoreductase) catalyzes NADH oxidation, O2 reduction to superoxide, and the reduction of several ‘artificial’ electron acceptors. Here, we show that the positively-charged electron acceptors paraquat and hexaammineruthenium(III) react with the nucleotide-bound reduced flavin in complex I, by an unusual ternary mechanism. NADH, ATP, ADP and ADP-ribose stimulate the reactions, indicating that the positively-charged acceptors interact with their negatively-charged phosphates. Our mechanism for paraquat reduction defines a new mechanism for superoxide production by complex I (by redox cycling); in contrast to direct O2 reduction the rate is stimulated, not inhibited, by high NADH concentrations

Penrose Diagram for a Transient Black Hole

A Penrose diagram is constructed for a spatially coherent black hole that smoothly begins an accretion, then excretes symmetrically as measured by a distant observer, with the initial and final states described by a metric of Minkowski form. Coordinate curves on the diagram are computationally derived. Causal relationships between space-time regions are briefly discussed. The life cycle of the black hole demonstrably leaves asymptotic observers in an unaltered Minkowski space-time of uniform conformal scale.Comment: 14 pages, 9 figures, spelling correction

A Beginner’s Guide to Thermodynamic Modelling of [FeFe] Hydrogenase

[FeFe] hydrogenases, which are considered the most active naturally occurring catalysts for hydrogen oxidation and proton reduction, are extensively studied as models to learn the important features for efficient H2 conversion catalysis. Using infrared spectroscopy as a selective probe, the redox behaviour of the active site H-cluster is routinely modelled with thermodynamic schemes based on the Nernst equation for determining thermodynamic parameters, such as redox midpoint potentials and pKa values. Here, the thermodynamic models usually applied to [FeFe] hydrogenases are introduced and discussed in a pedagogic fashion and their applicability to additional metalloenzymes and molecular catalysts is also addressed

An Observational Test of Holographic Inflation

Observational consequences of inflationary cosmology in the holographic dual of the Randall-Sundrum type II braneworld scenario, as motivated by the AdS/CFT correspondence, are investigated. High energy corrections to the standard four-dimensional Friedmann equation induce a corresponding modification to the form of the single-field inflationary consistency equation based on Einstein gravity. The degree of departure from the standard expression is determined by the ratio, r, of the primordial tensor and scalar perturbation amplitudes and the coefficient, c, of the conformal anomaly in the dual gauge theory. It is found that a necessary condition for detecting such a correction with the next generation of cosmic microwave background (CMB) polarization experiments is that r >= 0.06. The bound is tightened to r > 0.3 for values of the central charge that are compatible with known compactifications of type IIB string theory as parametrized in terms of F-theory compactification on Calabi-Yau four-folds. This is close to the present upper bounds inferred from combined observations of the CMB anisotropy power spectrum and high redshift surveys. We conclude that if such modifications to the inflationary consistency equation are to be observable, the gravitational wave background should be detected in the near future. A further consequence of the non-standard dynamics at high energies is that the initial state of the universe is a quiescent singularity with a finite density and pressure.Comment: 7 pages, 2 figures. Uses RevTeX4 LaTeX clas

Structural insight on the mechanism of an electron-bifurcating [FeFe] hydrogenase.

Electron-bifurcation is a fundamental energy conservation mechanism in nature in which two electrons from an intermediate potential electron donor are split so that one is sent along a high potential pathway to a high potential acceptor and the other is sent along a low potential pathway to a low potential acceptor. This process allows endergonic reactions to be driven by exergonic ones and is an alternative, less recognised, mechanism of energy coupling to the well-known chemiosmotic principle. The electron-bifurcating [FeFe] hydrogenase from Thermotoga maritima (HydABC) requires both NADH and ferredoxin to reduce protons generating hydrogen. The mechanism of electron-bifurcation in HydABC remains enigmatic in spite of intense research efforts over the last few years. Structural information may provide the basis for a better understanding of spectroscopic and functional information. Here, we present a 2.3 Å electron cryo-microscopy structure of HydABC. The structure shows a heterododecamer composed of two independent 'halves' each made of two strongly interacting HydABC heterotrimers connected via a [4Fe-4S] cluster. A central electron transfer pathway connects the active sites for NADH oxidation and for proton reduction. We identified two conformations of a flexible iron-sulfur cluster domain: a 'closed bridge' and an 'open bridge' conformation, where a Zn2+ site may act as a 'hinge' allowing domain movement. Based on these structural revelations, we propose a possible mechanism of electron-bifurcation in HydABC where the flavin mononucleotide serves a dual role as both the electron bifurcation center and as the NAD+ reduction/NADH oxidation site

Moduli Stabilization in Brane Gas Cosmology with Superpotentials

In the context of brane gas cosmology in superstring theory, we show why it is impossible to simultaneously stabilize the dilaton and the radion with a general gas of strings (including massless modes) and D-branes. Although this requires invoking a different mechanism to stabilize these moduli fields, we find that the brane gas can still play a crucial role in the early universe in assisting moduli stabilization. We show that a modest energy density of specific types of brane gas can solve the overshoot problem that typically afflicts potentials arising from gaugino condensation.Comment: minor changes to match the journal versio

Real time response on dS_3: the Topological AdS Black Hole and the Bubble

We study real time correlators in strongly coupled N=4 supersymmetric Yang-Mills theory on dS_3 x S^1, with antiperiodic boundary conditions for fermions on the circle. When the circle radius is larger than a critical value, the dual geometry is the so-called "topological AdS_5 black hole". Applying the Son- Starinets recipe in this background we compute retarded glueball propagators which exhibit an infinite set of poles yielding the quasinormal frequencies of the topological black hole. The imaginary parts of the propagators exhibit thermal effects associated with the Gibbons-Hawking temperature due to the cosmological horizon of the de Sitter boundary. We also obtain R-current correlators and find that after accounting for a small subtlety, the Son-Starinets prescription yields the retarded Green's functions. The correlators do not display diffusive behaviour at late times. Below the critical value of the circle radius, the topological black hole decays to the AdS_5 "bubble of nothing". Using a high frequency WKB approximation, we show that glueball correlators in this phase exhibit poles on the real axis. The tunnelling from the black hole to the bubble is interpreted as a hadronization transition.Comment: 52 pages, 11 figures, typos corrected, references adde

Redox tuning of the H-cluster by second coordination sphere amino acids in the sensory [FeFe] hydrogenase from Thermotoga maritima

[FeFe] hydrogenases are exceptionally active catalysts for the interconversion of molecular hydrogen with protons and electrons. Their active site, the H-cluster, is composed of a [4Fe-4S] cluster covalently linked to a unique [2Fe] subcluster. These enzymes have been extensively studied to understand how the protein environment tunes the properties of the Fe ions for efficient catalysis. The sensory [FeFe] hydrogenase (HydS) from Thermotoga maritima has low activity and displays a very positive redox potential for the [2Fe] subcluster compared to that of the highly active prototypical enzymes. Using site directed mutagenesis, we investigate how second coordination sphere interactions of the protein environment with the H-cluster in HydS influence the catalytic, spectroscopic and redox properties of the H-cluster. In particular, mutation of the non-conserved serine 267, situated between the [4Fe-4S] and [2Fe] subclusters, to methionine (conserved in prototypical catalytic enzymes) gave a dramatic decrease in activity. Infra-red (IR) spectroelectrochemistry revealed a 50 mV lower redox potential for the [4Fe-4S] subcluster in the S267M variant. We speculate that this serine forms a hydrogen bond to the [4Fe-4S] subcluster, increasing its redox potential. These results demonstrate the importance of the secondary coordination sphere in tuning the catalytic properties of the H-cluster in [FeFe] hydrogenases and reveal a particularly important role for amino acids interacting with the [4Fe-4S] subcluster

Time-Resolved Infrared Spectroscopy Reveals the pH-Independence of the First Electron Transfer Step in the [FeFe] Hydrogenase Catalytic Cycle.

[FeFe] hydrogenases are highly active catalysts for hydrogen conversion. Their active site has two components: a [4Fe-4S] electron relay covalently attached to the H2 binding site and a diiron cluster ligated by CO, CN-, and 2-azapropane-1,3-dithiolate (ADT) ligands. Reduction of the [4Fe-4S] site was proposed to be coupled with protonation of one of its cysteine ligands. Here, we used time-resolved infrared (TRIR) spectroscopy on the [FeFe] hydrogenase from Chlamydomonas reinhardtii (CrHydA1) containing a propane-1,3-dithiolate (PDT) ligand instead of the native ADT ligand. The PDT modification does not affect the electron transfer step to [4Fe-4S]H but prevents the enzyme from proceeding further through the catalytic cycle. We show that the rate of the first electron transfer step is independent of the pH, supporting a simple electron transfer rather than a proton-coupled event. These results have important implications for our understanding of the catalytic mechanism of [FeFe] hydrogenases and highlight the utility of TRIR

The Nonphysiological Reductant Sodium Dithionite and [FeFe] Hydrogenase: Influence on the Enzyme Mechanism.

[FeFe] hydrogenases are highly active enzymes for interconverting protons and electrons with hydrogen (H2). Their active site H-cluster is formed of a canonical [4Fe-4S] cluster ([4Fe-4S]H) covalently attached to a unique [2Fe] subcluster ([2Fe]H), where both sites are redox active. Heterolytic splitting and formation of H2 takes place at [2Fe]H, while [4Fe-4S]H stores electrons. The detailed catalytic mechanism of these enzymes is under intense investigation, with two dominant models existing in the literature. In one model, an alternative form of the active oxidized state Hox, named HoxH, which forms at low pH in the presence of the nonphysiological reductant sodium dithionite (NaDT), is believed to play a crucial role. HoxH was previously suggested to have a protonated [4Fe-4S]H. Here, we show that HoxH forms by simple addition of sodium sulfite (Na2SO3, the dominant oxidation product of NaDT) at low pH. The low pH requirement indicates that sulfur dioxide (SO2) is the species involved. Spectroscopy supports binding at or near [4Fe-4S]H, causing its redox potential to increase by ∼60 mV. This potential shift detunes the redox potentials of the subclusters of the H-cluster, lowering activity, as shown in protein film electrochemistry (PFE). Together, these results indicate that HoxH and its one-electron reduced counterpart Hred′H are artifacts of using a nonphysiological reductant, and not crucial catalytic intermediates. We propose renaming these states as the "dithionite (DT) inhibited"states Hox-DTi and Hred-DTi. The broader potential implications of using a nonphysiological reductant in spectroscopic and mechanistic studies of enzymes are highlighted