Fast voltammetric studies of the kinetics and energetics of coupled electron-transfer reactions in proteins

A wealth of information on the reactions of redox-active sites in proteins can be obtained by voltammetric studies in which the protein sample is arranged as a layer on an electrode surface. By carrying out cyclic voltammetry over a wide range of scan rates and exploiting the ability to poise or pulse the electrode potential between cycles, data are obtained that are conveniently (albeit simplistically) analysed in terms of plots of peak potentials against scan rate. A simple reversible electron-transfer process gives rise to a 'trumpet'-shaped plot because the oxidation and reduction peaks separate increasingly at high scan rate; the electrochemical kinetics are then determined by fitting to Butler-Volmer or Marcus models. Much more interesting though are the ways in which this 'trumpet plot' is altered, often dramatically, when electron transfer is coupled to biologically important processes such as proton transfer, ligand exchange, or a change in conformation. It is then possible to derive particularly detailed information on the kinetics, energetics and mechanism of reactions that may not revealed clearly or even at all by other methods. In order to interpret the voltammetry of coupled systems, it is important to be able to define 'ideal behaviour' for systems that are expected to show simple and uncoupled electron transfer. Accordingly, this paper describes results we have obtained for several proteins that are expected to show such behaviour, and compares these results with theoretical predictions.     

Fast, long-range electron-transfer reactions of a "blue" copper protein coupled non-covalently to an electrode through a stilbenyl thiolate monolayer

A self-assembled monolayer (SAM), formed by the insitu saponification of a stilbenyl thioacetate on a gold electrode, yields fast electron transfer (ET)(the exchange rate at zero driving force exceeds 1600 s-1) with adsorbed molecules of the blue copper protein, azurin, over a distance exceeding 15 angstroms .     

Redox enzymes in tethered membranes

An electrode surface is presented that enables the characterization of redox-active membrane enzymes in a native-like environment. An ubiquinol oxidase from Escherichia coli, cytochrome bo(3) (cbo(3)), has been co-immobilized into tethered bilayer lipid membranes (tBLMs). The tBLM is formed on gold surfaces functionalized with cholesterol tethers which insert into the lower leaflet of the membrane. The planar membrane architecture is formed by self-assembly of proteoliposomes, and its structure is characterized by surface plasmon resonance (SPR), electrochemical impedance spectroscopy (EIS), and tapping-mode atomic force microscopy (TM-AFM). The functionality of cbo(3) is investigated by cyclic voltammetry (CV) and is confirmed by the catalytic reduction of oxygen. Interfacial electron transfer to cbo(3) is mediated by the membrane-localized ubiquinol-8, the physiological electron donor of cbo(3). Enzyme coverages observed with TM-AFM and CV coincide (2-8.5 fmol.cm(-)(2)), indicating that most-if not all-cbo(3) on the surface is catalytically active and thus retains its integrity during immobilization.     

New99mTc-diiodine substituted IDA derivative (DIIODIDA) for hepatobiliary imaging

A new diiodine substituted IDA derivative, 2,4-diiodine-6-methyl IDA (DIIODIDA) was synthesized and labeled with^99mTc. It was established that^99mTc-DIIODIDA had high radiochemical purity. Biodistribution and influence of bilirubin on^99mTc-DIIODIDA biokinetics has been studied in rats and compared to the corresponding results for^99mTc-SOLCOIODIDA. Related to^99mTc-SOLCOIODIDA,^99mTc-DIIODIDA has much better biliary exretion (55.18 versus 43.63%). No change of^99mTc-DIIODIDA biokinetics, under influence of bilirubin was noticed. Biliary excretion of^99mTc-SOLCOIODIDA has been reduced for about 60%. The protein binding of^99mTc-DIIODIDA and^99mTc-SOLCOIODIDA were also determined, using in vitro method of precipitation. These results showed that^99mTc-DIIODIDA hepatobiliary imaging agent is superior to the presently used^99mTc-monoiodine IDA derivatives.     

一般n阶特征量泛函的Euler-Lagrange方程及与定量因果原理、相对性原理和广义牛顿三定律的统一

本文获得了有各种相互作用的一般n阶特征量泛函,其耦合系数反映了不同特征量泛函之间的耦合强度.依据定量因果原理,导出了一般n阶特征量泛函的变分原理,获得了一般n阶特征量泛函的Euler-Lagrange方程,它的不同系数可拟合不同的物理现实,如从线性到任意n阶非线性物理系统,使复杂难解的任意n阶非线性物理系统变得具体可解.并获得了该对称变换下不变的m个的守恒量,以及它们之间的关系和统一描述.依据定量因果原理导出了相对性原理,证明了绝对加速参考系、牵连参考系和相对参考系的力都有来自加速度和质量变化的贡献.利用定量因果原理自然导出了广义牛顿第一定律和广义牛顿第二定律,而且还导出了一个新定律,即广义牛顿第三定律,亦即平移不变性系统合力为零定理.进而将研究结论应用于对银河系的修正引力势、分子势、夸克禁闭势等,且其结果与物理实验一致.     

On relative stability of linear stationary feedback control systems with time delay

A method is presented whereby the absolute and relative stabilityof linear control systems containing transport lag can be determined.As a result feedback systems with variable time delay and loopgain, may be investigated in straightforward manner.     

Electron-transfer mechanisms through biological redox chains in multicenter enzymes

A new approach for studying intramolecular electron transfer in multicenter enzymes is described. Two fumarate reductases, adsorbed on an electrode in a fully active state, have been studied using square-wave voltammetry as a kinetic method to probe the mechanism of the long-range electron transfer to and from the buried active site. Flavocytochrome c(3) (Fcc(3)), the globular fumarate reductase from Shewanella frigidimarina, and the soluble subcomplex of the membrane-bound fumarate reductase of Escherichia coli (FrdAB) each contain an active site FAD that is redox-connected to the surface by a chain of hemes or Fe-S clusters, respectively. Using square-wave voltammetry with large amplitudes, we have measured the electron-transfer kinetics of the FAD cofactor as a function of overpotential. The results were modeled in terms of the FAD group receiving or donating electrons either via a direct mechanism or one involving hopping via the redox chain. The FrdAB kinetics could be described by both models, while the Fcc(3) data could only be fit on the basis of a direct electron-transfer mechanism. This raises the likelihood that electron transfer can occur via a superexchange mechanism utilizing the heme groups to enhance electronic coupling. Finally, the FrdAB data show, in contrast to Fcc(3), that the maximum ET rate at high overpotential is related to the turnover number for FrdAB measured previously so that electron transfer is the limiting step during catalysis.