An energy coupling factor transporter of Streptococcus sanguinis impacts antibiotic susceptibility as well as metal and membrane homeostasis.

Streptococcus sanguinis is a prevalent member of human microbiome capable of acting as a causative agent of oral and respiratory infections. S. sanguinis competitive success within the infection niche is dependent on acquisition of metal ions and vitamins. Among the systems that bacteria use for micronutrient uptake is the energy coupling factor (ECF) transporter system EcfAAT. Here we describe physiological changes arising from EcfAAT transporter disruption. We found that EcfAAT contributes to S. sanguinis antibiotic sensitivity as well as metal and membrane homeostasis. Specifically, our work found that disruption of EcfAAT results in increased polymyxin susceptibility. We performed assessment of cell-associated metal content and found depletion of iron, magnesium, and manganese. Furthermore, membrane composition analysis revealed significant enrichment in unsaturated fatty acid species resulting in increased membrane fluidity. Our results demonstrate how disruption of a single EcfAAT transporter can have broad consequences on bacterial cell homeostasis. ECF transporters are of interest within the context of infection biology in bacterial species other than streptococci, hence work described here will further the understanding of how micronutrient uptake systems contribute to bacterial pathogenesis.

Confined Element Distribution with Structure-Driven Energy Coupling for Enhanced Prussian Blue Analogue Cathode.

The structural failure of Na2Mn[Fe(CN)6] could not be alleviated with traditional modification strategies through the adjustable composition property of Prussian blue analogues (PBAs), considering that the accumulation and release of stress derived from the MnN6 octahedrons are unilaterally restrained. Herein, a novel application of adjustable composition property, through constructing a coordination competition relationship between chelators and [Fe(CN)6]4- to directionally tune the enrichment of elements, is proposed to restrain structural degradation and induce unconventional energy coupling phenomenon. The non-uniform distribution of elements at the M1 site of PBAs (NFM-PB) is manipulated by the sequentially precipitated Ni, Fe, and Mn according to the Irving-William order. Electrochemically active Fe is operated to accompany Mn, and zero-strain Ni is modulated to enrich at the surface, synergistically mitigating with the enrichment and release of stress and then significantly improving the structural stability. Furthermore, unconventional energy coupling effect, a fusion of the electrochemical behavior between FeLS and MnHS, is triggered by the confined element distribution, leading to the enhanced electrochemical stability and anti-polarization ability. Consequently, the NFM-PB demonstrates superior rate performance and cycling stability. These findings further exploit potentialities of the adjustable composition property and provide new insights into the component design engineering for advanced PBAs.

Size matters in metabolic scaling: Critical role of the thermodynamic efficiency of ATP synthesis and its dependence on mitochondrial H+ leak across mammalian species.

In this last article of the trilogy, the unified biothermokinetic theory of ATP synthesis developed in the previous two papers is applied to a major problem in comparative physiology, biochemistry, and ecology-that of metabolic scaling as a function of body mass across species. A clear distinction is made between intraspecific and interspecific relationships in energy metabolism, clearing up confusion that had existed from the very beginning since Kleiber first proposed his mouse-to-elephant rule almost a century ago. It is shown that the overall mass exponent of basal/standard metabolic rate in the allometric relationship [Formula: see text] is composed of two parts, one emerging from the relative intraspecific constancy of the slope (b), and the other (b') arising from the interspecific variation of the mass coefficient, a(M) with body size. Quantitative analysis is shown to reveal the hidden underlying relationship followed by the interspecific mass coefficient, a(M)=P0M0.10, and a universal value of P0=3.23 watts, W is derived from empirical data on mammals from mouse to cattle. The above relationship is shown to be understood only within an evolutionary biological context, and provides a physiological explanation for Cope's rule. The analysis also helps in fundamentally understanding how variability and a diversity of scaling exponents arises in allometric relations in biology and ecology. Next, a molecular-level understanding of the scaling of metabolism across mammalian species is shown to be obtained by consideration of the thermodynamic efficiency of ATP synthesis η, taking mitochondrial proton leak as a major determinant of basal metabolic rate in biosystems. An iterative solution is obtained by solving the mathematical equations of the biothermokinetic ATP theory, and the key thermodynamic parameters, e.g. the degree of coupling q, the operative P/O ratio, and the metabolic efficiency of ATP synthesis η are quantitatively evaluated for mammals from rat to cattle. Increases in η (by ∼15%) over a 2000-fold body size range from rat to cattle, primarily arising from an ∼3-fold decrease in the mitochondrial H+ leak rate are quantified by the unified ATP theory. Biochemical and mechanistic consequences for the interpretation of basal metabolism, and the various molecular implications arising are discussed in detail. The results are extended to maximum metabolic rate, and interpreted mathematically as a limiting case of the general ATP theory. The limitations of the analysis are pointed out. In sum, a comprehensive quantitative analysis based on the unified biothermokinetic theory of ATP synthesis is shown to solve a central problem in biology, physiology, and ecology on the scaling of energy metabolism with body size.

Thermodynamic analysis of energy coupling by determination of the Onsager phenomenological coefficients for a 3×3 system of coupled chemical reactions and transport in ATP synthesis and its mechanistic implications.

The nonequilibrium coupled processes of oxidation and ATP synthesis in the fundamental process of oxidative phosphorylation (OXPHOS) are of vital importance in biosystems. These coupled chemical reaction and transport bioenergetic processes using the OXPHOS pathway meet >90% of the ATP demand in aerobic systems. On the basis of experimentally determined thermodynamic OXPHOS flux-force relationships and biochemical data for the ternary system of oxidation, ion transport, and ATP synthesis, the Onsager phenomenological coefficients have been computed, including an estimate of error. A new biothermokinetic theory of energy coupling has been formulated and on its basis the thermodynamic parameters, such as the overall degree of coupling, q and the phenomenological stoichiometry, Z of the coupled system have been evaluated. The amount of ATP produced per oxygen consumed, i.e. the actual, operating P/O ratio in the biosystem, the thermodynamic efficiency of the coupled reactions, η, and the Gibbs free energy dissipation, Φ have been calculated and shown to be in agreement with experimental data. At the concentration gradients of ADP and ATP prevailing under state 3 physiological conditions of OXPHOS that yield Vmax rates of ATP synthesis, a maximum in Φ of ∼0.5J(hmgprotein)-1, corresponding to a thermodynamic efficiency of ∼60% for oxidation on succinate, has been obtained. Novel mechanistic insights arising from the above have been discussed. This is the first report of a 3 × 3 system of coupled chemical reactions with transport in a biological context in which the phenomenological coefficients have been evaluated from experimental data.

The Warburg Effect Reinterpreted 100 yr on: A First-Principles Stoichiometric Analysis and Interpretation from the Perspective of ATP Metabolism in Cancer Cells.

The Warburg Effect is a longstanding enigma in cancer biology. Despite the passage of 100 yr since its discovery, and the accumulation of a vast body of research on the subject, no convincing biochemical explanation has been given for the original observations of aerobic glycolysis in cancer cell metabolism. Here, we have worked out a first-principles quantitative analysis of the problem from the principles of stoichiometry and available electron balance. The results have been interpreted using Nath's unified theory of energy coupling and adenosine triphosphate (ATP) synthesis, and the original data of Warburg and colleagues have been analyzed from this new perspective. Use of the biomass yield based on ATP per unit substrate consumed, [Formula: see text], or the Nath-Warburg number, NaWa has been shown to excellently model the original data on the Warburg Effect with very small standard deviation values, and without employing additional fitted or adjustable parameters. Based on the results of the quantitative analysis, a novel conservative mechanism of synthesis, utilization, and recycling of ATP and other key metabolites (eg, lactate) is proposed. The mechanism offers fresh insights into metabolic symbiosis and coupling within and/or among proliferating cells. The fundamental understanding gained using our approach should help in catalyzing the development of more efficient metabolism-targeting anticancer drugs.

Volumetric Modification of Transparent Materials with Two-Color Laser Irradiation: Insight from Numerical Modeling.

Traditionally, single-color laser beams are used for material processing and modifications of optical, mechanical, conductive, and thermal properties of different materials. So far, there are a limited number of studies about the dual-wavelength laser irradiation of materials, which, however, indicate a strong enhancement in laser energy coupling to solid targets. Here, a theoretical study is reported that aimed at exploring the volumetric excitation of fused silica with dual-wavelength (800 nm and 400 nm) ultrashort laser pulses focused on the material's bulk. Numerical simulations are based on Maxwell's equations, accounting for the generation of conduction electrons, their hydrodynamic motion in the laser field, and trapping into an excitonic state. It is shown that, by properly choosing the energies of the two laser harmonics successively coupling with the material, it is possible to strongly enhance the laser energy absorption as compared to the pulses of a single wavelength with the same total energy. Laser energy absorption strongly depends on the sequence of applied wavelengths, so that the shorter wavelength pre-irradiation can yield a dramatic effect on laser excitation by the following longer-wavelength pulse. The predictions of this study can open a new route for enhancing and controlling the highly localized absorption of laser energy inside transparent materials for optoelectronic and photonic applications.

Coupling and biological free-energy transduction processes as a bridge between physics and life: Molecular-level instantiation of Ervin Bauer's pioneering concepts in biological thermodynamics.

The nonequilibrium coupled processes of oxidation and ATP synthesis in the biological process of oxidative phosphorylation (OXPHOS) are fundamental to all life on our planet. These steady-state energy transduction processes ‒ coupled by proton and anion/counter-cation concentration gradients in the OXPHOS pathway ‒ generate ∼95 % of the ATP requirement of aerobic systems for cellular function. The rapid energy cycling and homeostasis of metabolites involved in this coupling are shown to be responsible for maintenance and regulation of stable nonequilibrium states, the latter first postulated in pioneering biothermodynamics work by Ervin Bauer between 1920 and 1935. How exactly does this occur? This is shown to be answered by molecular considerations arising from Nath's torsional mechanism of ATP synthesis and two-ion theory of energy coupling developed in 25 years of research work on the subject. A fresh analysis of the biological thermodynamics of coupling that goes beyond the previous work of Stucki and others and shows how the system functions at the molecular level has been carried out. Thermodynamic parameters, such as the overall degree of coupling, q of the coupled system are evaluated for the state 4 to state 3 transition in animal mitochondria with succinate as substrate. The actual or operative P to O ratio, the efficiency of the coupled reactions, η, and the Gibbs energy dissipation, Φ have been calculated and shown to be in good agreement with experimental data. Novel mechanistic insights arising from the above have been discussed. A fourth law/principle of thermodynamics is formulated for a sub-class of physical and biological systems. The critical importance of constraints and time-varying boundary conditions for function and regulation is discussed in detail. Dynamic internal structural changes essential for torsional energy storage within the γ-subunit in a single molecule of the FOF1-ATP synthase and its transduction have been highlighted. These results provide a molecular-level instantiation of Ervin Bauer's pioneering concepts in biological thermodynamics.

Elucidating Events within the Black Box of Enzyme Catalysis in Energy Metabolism: Insights into the Molecular Mechanism of ATP Hydrolysis by F1-ATPase.

Oxygen exchange reactions occurring at ß-catalytic sites of the FOF1-ATP synthase/F1-ATPase imprint a unique record of molecular events during the catalytic cycle of ATP synthesis/hydrolysis. This work presents a new theory of oxygen exchange and tests it on oxygen exchange data recorded on ATP hydrolysis by mitochondrial F1-ATPase (MF1). The apparent rate constant of oxygen exchange governing the intermediate Pi-HOH exchange accompanying ATP hydrolysis is determined by kinetic analysis over a ~50,000-fold range of substrate ATP concentration (0.1-5000 µM) and a corresponding ~200-fold range of reaction velocity (3.5-650 [moles of Pi/{moles of F1-ATPase}-1 s-1]). Isotopomer distributions of [18O]Pi species containing 0, 1, 2, and 3 labeled oxygen atoms predicted by the theory have been quantified and shown to be in perfect agreement with the experimental distributions over the entire range of medium ATP concentrations without employing adjustable parameters. A novel molecular mechanism of steady-state multisite ATP hydrolysis by the F1-ATPase has been proposed. Our results show that steady-state ATP hydrolysis by F1-ATPase occurs with all three sites occupied by Mg-nucleotide. The various implications arising from models of energy coupling in ATP synthesis/hydrolysis by the ATP synthase/F1-ATPase have been discussed. Current models of ATP hydrolysis by F1-ATPase, including those postulated from single-molecule data, are shown to be effectively bisite models that contradict the data. The trisite catalysis formulated by Nath's torsional mechanism of energy transduction and ATP synthesis/hydrolysis since its first appearance 25 years ago is shown to be in better accord with the experimental record. The total biochemical information on ATP hydrolysis is integrated into a consistent model by the torsional mechanism of ATP synthesis/hydrolysis and shown to elucidate the elementary chemical and mechanical events within the black box of enzyme catalysis in energy metabolism by F1-ATPase.

Beyond binding change: the molecular mechanism of ATP hydrolysis by F1-ATPase and its biochemical consequences.

F1-ATPase is a universal multisubunit enzyme and the smallest-known motor that, fueled by the process of ATP hydrolysis, rotates in 120o steps. A central question is how the elementary chemical steps occurring in the three catalytic sites are coupled to the mechanical rotation. Here, we performed cold chase promotion experiments and measured the rates and extents of hydrolysis of preloaded bound ATP and promoter ATP bound in the catalytic sites. We found that rotation was caused by the electrostatic free energy change associated with the ATP cleavage reaction followed by Pi release. The combination of these two processes occurs sequentially in two different catalytic sites on the enzyme, thereby driving the two rotational sub-steps of the 120o rotation. The mechanistic implications of this finding are discussed based on the overall energy balance of the system. General principles of free energy transduction are formulated, and their important physical and biochemical consequences are analyzed. In particular, how exactly ATP performs useful external work in biomolecular systems is discussed. A molecular mechanism of steady-state, trisite ATP hydrolysis by F1-ATPase, consistent with physical laws and principles and the consolidated body of available biochemical information, is developed. Taken together with previous results, this mechanism essentially completes the coupling scheme. Discrete snapshots seen in high-resolution X-ray structures are assigned to specific intermediate stages in the 120o hydrolysis cycle, and reasons for the necessity of these conformations are readily understood. The major roles played by the "minor" subunits of ATP synthase in enabling physiological energy coupling and catalysis, first predicted by Nath's torsional mechanism of energy transduction and ATP synthesis 25 years ago, are now revealed with great clarity. The working of nine-stepped (bMF1, hMF1), six-stepped (TF1, EF1), and three-stepped (PdF1) F1 motors and of the α3ß3γ subcomplex of F1 is explained by the same unified mechanism without invoking additional assumptions or postulating different mechanochemical coupling schemes. Some novel predictions of the unified theory on the mode of action of F1 inhibitors, such as sodium azide, of great pharmaceutical importance, and on more exotic artificial or hybrid/chimera F1 motors have been made and analyzed mathematically. The detailed ATP hydrolysis cycle for the enzyme as a whole is shown to provide a biochemical basis for a theory of "unisite" and steady-state multisite catalysis by F1-ATPase that had remained elusive for a very long time. The theory is supported by a probability-based calculation of enzyme species distributions and analysis of catalytic site occupancies by Mg-nucleotides and the activity of F1-ATPase. A new concept of energy coupling in ATP synthesis/hydrolysis based on fundamental ligand substitution chemistry has been advanced, which offers a deeper understanding, elucidates enzyme activation and catalysis in a better way, and provides a unified molecular explanation of elementary chemical events occurring at enzyme catalytic sites. As such, these developments take us beyond binding change mechanisms of ATP synthesis/hydrolysis proposed for oxidative phosphorylation and photophosphorylation in bioenergetics.

Structure, Assembly, and Function of Flagella Responsible for Bacterial Locomotion.

Many motile bacteria use flagella for locomotion under a variety of environmental conditions. Because bacterial flagella are under the control of sensory signal transduction pathways, each cell is able to autonomously control its flagellum-driven locomotion and move to an environment favorable for survival. The flagellum of Salmonella enterica serovar Typhimurium is a supramolecular assembly consisting of at least three distinct functional parts: a basal body that acts as a bidirectional rotary motor together with multiple force generators, each of which serves as a transmembrane proton channel to couple the proton flow through the channel with torque generation; a filament that functions as a helical propeller that produces propulsion; and a hook that works as a universal joint that transmits the torque produced by the rotary motor to the helical propeller. At the base of the flagellum is a type III secretion system that transports flagellar structural subunits from the cytoplasm to the distal end of the growing flagellar structure, where assembly takes place. In recent years, high-resolution cryo-electron microscopy (cryoEM) image analysis has revealed the overall structure of the flagellum, and this structural information has made it possible to discuss flagellar assembly and function at the atomic level. In this article, we describe what is known about the structure, assembly, and function of Salmonella flagella.

Active Site Engineering on Plasmonic Nanostructures for Efficient Photocatalysis.

Plasmonic nanostructures have shown immense potential in photocatalysis because of their distinct photochemical properties associated with tunable photoresponses and strong light-matter interactions. The introduction of highly active sites is essential to fully exploit the potential of plasmonic nanostructures in photocatalysis, considering the inferior intrinsic activities of typical plasmonic metals. This review focuses on active site-engineered plasmonic nanostructures with enhanced photocatalytic performance, wherein the active sites are classified into four types (i.e., metallic sites, defect sites, ligand-grafted sites, and interface sites). The synergy between active sites and plasmonic nanostructures in photocatalysis is discussed in detail after briefly introducing the material synthesis and characterization methods. Active sites can promote the coupling of solar energy harvested by plasmonic metal to catalytic reactions in the form of local electromagnetic fields, hot carriers, and photothermal heating. Moreover, efficient energy coupling potentially regulates the reaction pathway by facilitating the excited state formation of reactants, changing the status of active sites, and creating additional active sites using photoexcited plasmonic metals. Afterward, the application of active site-engineered plasmonic nanostructures in emerging photocatalytic reactions is summarized. Finally, a summary and perspective of the existing challenges and future opportunities are presented. This review aims to deliver some insights into plasmonic photocatalysis from the perspective of active sites, expediting the discovery of high-performance plasmonic photocatalysts.

Distinct Energy-Coupling Factor Transporter Subunits Enable Flavin Acquisition and Extracytosolic Trafficking for Extracellular Electron Transfer in Listeria monocytogenes.

A variety of electron transfer mechanisms link bacterial cytosolic electron pools with functionally diverse redox activities in the cell envelope and extracellular space. In Listeria monocytogenes, the ApbE-like enzyme FmnB catalyzes extracytosolic protein flavinylation, covalently linking a flavin cofactor to proteins that transfer electrons to extracellular acceptors. L. monocytogenes uses an energy-coupling factor (ECF) transporter complex that contains distinct substrate-binding, transmembrane, ATPase A, and ATPase A' subunits (RibU, EcfT, EcfA, and EcfA') to import environmental flavins, but the basis of extracytosolic flavin trafficking for FmnB flavinylation remains poorly defined. In this study, we show that the EetB and FmnA proteins are related to ECF transporter substrate-binding and transmembrane subunits, respectively, and are essential for exporting flavins from the cytosol for flavinylation. Comparisons of the flavin import versus export capabilities of L. monocytogenes strains lacking different ECF transporter subunits demonstrate a strict directionality of substrate-binding subunit transport but partial functional redundancy of transmembrane and ATPase subunits. Based on these results, we propose that ECF transporter complexes with different subunit compositions execute directional flavin import/export through a broadly conserved mechanism. Finally, we present genomic context analyses that show that related ECF exporter genes are distributed across members of the phylum Firmicutes and frequently colocalize with genes encoding flavinylated extracytosolic proteins. These findings clarify the basis of ECF transporter export and extracytosolic flavin cofactor trafficking in Firmicutes. IMPORTANCE Bacteria import vitamins and other essential compounds from their surroundings but also traffic related compounds from the cytosol to the cell envelope where they serve various functions. Studying the foodborne pathogen Listeria monocytogenes, we find that the modular use of subunits from a prominent class of bacterial transporters enables the import of environmental vitamin B2 cofactors and the extracytosolic trafficking of a vitamin B2-derived cofactor that facilitates redox reactions in the cell envelope. These studies clarify the basis of bidirectional small-molecule transport across the cytoplasmic membrane and the assembly of redox-active proteins within the cell envelope and extracellular space.

Fluorescence Resonance Energy Transfer (FRET) as a Spectroscopic Ruler for the Investigation of Protein Induced Lipid Membrane Curvature: Bacteriorhodopsin and Bacteriorhodopsin Analogs in Model Lipid Membranes.

Bacteriorhodopsin (bR) is a light-driven proton pump existing in the purple membranes (PM) of Halobacterium salinarum. The effects associated with changes in proton distribution (proton gradient, membrane electric potential) play a key role in ATPase stimulation. However, how the bioenergetic modulus (bR-PM-ATPase) functions remains unclear. One can find indications that hydrophobic matching and the curvature of the lipid membrane may form a functional link between bR and ATPase. To verify whether an interaction between bR and lipids can lead to curvature of the lipid membrane, a spectroscopic ruler, that is, a fluorescence resonance energy transfer (FRET) tool, was used. The distances from fluorescent lipid probes [octadecyl rhodamine B chloride (RhB), 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI), 16-(9-anthroyloxy) palmitic acid (16AP), and hydrophobic probe 1,6-diphenyl-1,3,5-hexatriene (DPH), to the retinal chromophore of bR incorporated into phospholipid vesicles, were measured. The incorporation of retinal analogues with changed shape and/or altered electronic properties into the binding site of a bR or bR mutant were used to strengthen the feedback between the protein surrounding and chromophore. The experiments were performed with wild-type and D96N-mutated bR carrying retinal or 14-(12-,10-, 13,14-bi-) fluororetinal. As far as it is known, this is the first time that results obtained by the FRET method show that bR can induce a change in lipid structure interpreted as hydrophobically induced curving of the lipid membrane. Evidence was provided that the chromophore contributed to this effect. The extent of contribution was dependent on the chromophore structure in close vicinity to the place of its link with opsin. The implications of these findings for bR-PM-ATPase module functioning are also discussed.

The Need for Consistency with Physical Laws and Logic in Choosing Between Competing Molecular Mechanisms in Biological Processes: A Case Study in Modeling ATP Synthesis.

Traditionally, proposed molecular mechanisms of fundamental biological processes have been tested against experiment. However, owing to a plethora of reasons-difficulty in designing, carrying out, and interpreting key experiments, use of different experimental models and systems, conduct of studies under widely varying experimental conditions, fineness in distinctions between competing mechanisms, complexity of the scientific issues, and the resistance of some scientists to discoveries that are contrary to popularly held beliefs-this has not solved the problem despite decades of work in the field/s. The author would like to prescribe an alternative way: that of testing competing models/mechanisms for their adherence to scientific laws and principles, and checking for errors in logic. Such tests are fairly commonly carried out in the mathematics, physics, and engineering literature. Further, reported experimental measurements should not be smaller than minimum detectable values for the measurement technique employed and should truly reflect function of the actual system without inapplicable extrapolation. Progress in the biological fields would be greatly accelerated, and considerable scientific acrimony avoided by adopting this approach. Some examples from the fundamental field of ATP synthesis in oxidative phosphorylation (OXPHOS) have been reviewed that also serve to illustrate the approach. The approach has never let the author down in his 35-yr-long experience on biological mechanisms. This change in thinking should lead to a considerable saving of both time and resources, help channel research efforts toward solution of the right problems, and hopefully provide new vistas to a younger generation of open-minded biological scientists.

Construction of magnetic nanochains to achieve magnetic energy coupling in scaffold.

BACKGROUND: Fe3O4 nanoparticles are highly desired for constructing endogenous magnetic microenvironment in scaffold to accelerate bone regeneration due to their superior magnetism. However, their random arrangement easily leads to mutual consumption of magnetic poles, thereby weakening the magnetic stimulation effect. METHODS: In this study, magnetic nanochains are synthesized by magnetic-field-guided interface co-assembly of Fe3O4 nanoparticles. In detail, multiple Fe3O4 nanoparticles are aligned along the direction of magnetic force lines and are connected in series to form nanochain structures under an external magnetic field. Subsequently, the nanochain structures are covered and fixed by depositing a thin layer of silica (SiO2), and consequently forming linear magnetic nanochains (Fe3O4@SiO2). The Fe3O4@SiO2 nanochains are then incorporated into poly l-lactic acid (PLLA) scaffold prepared by selective laser sintering technology. RESULTS: The results show that the Fe3O4@SiO2 nanochains with unique core-shell structure are successfully constructed. Meanwhile, the orderly assembly of nanoparticles in the Fe3O4@SiO2 nanochains enable to form magnetic energy coupling and obtain a highly magnetic micro-field. The in vitro tests indicate that the PLLA/Fe3O4@SiO2 scaffolds exhibit superior capacity in enhancing cell activity, improving osteogenesis-related gene expressions, and inducing cell mineralization compared with PLLA and PLLA/Fe3O4 scaffolds. CONCLUSION: In short, the Fe3O4@SiO2 nanochains endow scaffolds with good magnetism and cytocompatibility, which have great potential in accelerating bone repair.

The Mechanism of Energy Coupling in H+/Na+-Pumping Membrane Pyrophosphatase-Possibilities and Probabilities.

Membrane pyrophosphatases (mPPases) found in plant vacuoles and some prokaryotes and protists are ancient cation pumps that couple pyrophosphate hydrolysis with the H+ and/or Na+ transport out of the cytoplasm. Because this function is reversible, mPPases play a role in maintaining the level of cytoplasmic pyrophosphate, a known regulator of numerous metabolic reactions. mPPases arouse interest because they are among the simplest membrane transporters and have no homologs among known ion pumps. Detailed phylogenetic studies have revealed various subtypes of mPPases and suggested their roles in the evolution of the "sodium" and "proton" bioenergetics. This treatise focuses on the mechanistic aspects of the transport reaction, namely, the coupling step, the role of the chemically produced proton, subunit cooperation, and the relationship between the proton and sodium ion transport. The available data identify H+-PPases as the first non-oxidoreductase pump with a "direct-coupling" mechanism, i.e., the transported proton is produced in the coupled chemical reaction. They also support a "billiard" hypothesis, which unifies the H+ and Na+ transport mechanisms in mPPase and, probably, other transporters.

Pre-steady-state kinetics and solvent isotope effects support the "billiard-type" transport mechanism in Na+ -translocating pyrophosphatase.

Membrane-bound pyrophosphatase (mPPase) found in microbes and plants is a membrane H+ pump that transports the H+ ion generated in coupled pyrophosphate hydrolysis out of the cytoplasm. Certain bacterial and archaeal mPPases can in parallel transport Na+ via a hypothetical "billiard-type" mechanism, also involving the hydrolysis-generated proton. Here, we present the functional evidence supporting this coupling mechanism. Rapid-quench and pulse-chase measurements with [32 P]pyrophosphate indicated that the chemical step (pyrophosphate hydrolysis) is rate-limiting in mPPase catalysis and is preceded by a fast isomerization of the enzyme-substrate complex. Na+ , whose binding is a prerequisite for the hydrolysis step, is not required for substrate binding. Replacement of H2 O with D2 O decreased the rates of pyrophosphate hydrolysis by both Na+ - and H+ -transporting bacterial mPPases, the effect being more significant than with a non-transporting soluble pyrophosphatase. We also show that the Na+ -pumping mPPase of Thermotoga maritima resembles other dimeric mPPases in demonstrating negative kinetic cooperativity and the requirement for general acid catalysis. The findings point to a crucial role for the hydrolysis-generated proton both in H+ -pumping and Na+ -pumping by mPPases.

Physical Basis of Multi-Energy Coupling-Driven Water Oxidation.

Hydrogen production by electrolyzing water is an important technique to store energy from renewables into chemical energy. Many efforts have been made to improve the energy conversion efficiency. In this review article, we mainly summarized the emerging ideas on water oxidation by multi-energy coupling. First, the physicochemical nature of electrolyzing water reaction is described. Then, we conceptually proposed the physical basis of energy coupling with a goal to maximize the energy conversion efficiency and showed the methods to achieve heat-electricity and magnetism-electricity coupling to drive water splitting. Finally, the material requirements for creating efficient energy coupling water splitting system were proposed. These new ideas unlock a big potential direction for developing multi-energy coupling hydrogen production devices to efficiently store the intermittent and fluctuating renewables.

Supercomplex supercomplexes: Raison d'etre and functional significance of supramolecular organization in oxidative phosphorylation.

Following structural determination by recent advances in electron cryomicroscopy, it is now well established that the respiratory Complexes I-IV in oxidative phosphorylation (OXPHOS) are organized into supercomplexes in the respirasome. Nonetheless, the reason for the existence of the OXPHOS supercomplexes and their functional role remains an enigma. Several hypotheses have been proposed for the existence of these supercomplex supercomplexes. A commonly-held view asserts that they enhance catalysis by substrate channeling. However, this - and other views - has been challenged based on structural and biophysical information. Hence, new ideas, concepts, and frameworks are needed. Here, a new model of energy transfer in OXPHOS is developed on the basis of biochemical data on the pure competitive inhibition of anionic substrates like succinate by the classical anionic uncouplers of OXPHOS (2,4-dinitrophenol, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone, and dicoumarol), and pharmacological data on the unique site-selective, energy-linked inhibition of energy conservation pathways in mitochondria induced by the guanidine derivatives. It is further found that uncouplers themselves are site-specific and exhibit differential selectivity and efficacy in reversing the inhibition caused by the Site 1/Complex I or Site 2/Complexes II-III-selective guanidine derivatives. These results lead to new vistas and sufficient complexity in the network of energy conservation pathways in the mitochondrial respiratory chain that necessitate discrete points of interaction with two classes of guanidine derivatives and uncoupling agents and thereby separate and distinct energy transfer pathways between Site 1 and Site 2 and the intermediate that energizes adenosine triphosphate (ATP) synthesis by Complex V. Interpretation based on Mitchell's single-ion chemiosmotic theory that postulates only a single energy pool is inadequate to rationalize the data and account for the required complexity. The above results and available information are shown to be explained by Nath's two-ion theory of energy coupling and ATP synthesis, involving coupled movement of succinate anions and protons, along with the requirement postulated by the theory for maintenance of homeostasis and ion translocation across the energy-transducing membrane of both succinate monoanions and succinate dianions by Complexes I-V in the OXPHOS supercomplexes. The new model of energy transfer in mitochondria is mapped onto the solved structures of the supercomplexes and integrated into a consistent model with the three-dimensional electron microscope computer tomography visualization of the internal structure of the cristae membranes in mammalian mitochondria. The model also offers valuable insights into diseased states induced in type 2 diabetes and especially in Alzheimer's and other neurodegenerative diseases that involve mitochondrial dysfunction.

Network representation and analysis of energy coupling mechanisms in cellular metabolism by a graph-theoretical approach.

Mechanisms coupling the chemical reactions of oxidation and ATP synthesis in cellular metabolism by the fundamental biological process of oxidative phosphorylation (OX PHOS) in mitochondria provide > 90% of the energy requirements in living organisms. Mathematical graph theory methods have been extensively used to characterize various metabolic, regulatory, and disease networks in biology. However, networks of energy coupling mechanisms in OX PHOS have not been represented and analyzed previously by these approaches. Here, the problem of biological energy coupling is translated into a graph-theoretical framework, and all possible coupling schemes between oxidation and ATP synthesis are represented as graphs connecting these processes by various intermediates or states. The problem is shown to be transformed into the hard problem of finding a Hamiltonian tour in the networks of possible constituent mechanisms, given the constraints of a cyclical nature of operation of enzymes and biological molecular machines. Accessible mathematical proofs of three theorems that guarantee sufficient conditions for the existence of a Hamiltonian cycle in simple graphs are provided. The results of the general theorems are applied to the set of possible coupling mechanisms in OX PHOS and shown to (1) unequivocally differentiate between the major theories and mechanisms of energy coupling, (2) greatly reduce the possibilities for detailed consideration, and (3) deduce the biologically selected mechanism using additional constraints from the cumulative experimental record. Finally, an algorithm is constructed to implement the graph-theoretical procedure. In summary, the enormous power and generality of mathematical theorems and approaches in graph theory are shown to help solve a fundamental problem in biology.