Proteins redox alterations

A variety of physical methods has been used to ascertain whether or not surface ruthenation alters the structure of a protein. UV-vis, CD, EPR, and resonance Raman spectroscopies have demonstrated that myoglobin [14, 18], cytochrome c [5, 16, 19, 21], and azurin [13] are not perturbed structurally by the attachment of a ruthenium complex to a surface histidine. The reduction potential of the metal redox center of a protein and its temperature dependence are indicators of protein structure as well. Cyclic voltammetry [5, 13], differential pulse polarography [14,21], and spectroelectrochemistry [12,14,22] are commonly used for the determination of the ruthenium and protein redox center potentials in modified proteins. 

Protons are translocated across the membrane by what is described as a proton pnmp . How does the pump operate The change in redox state experienced by the prosthetic gronps of the enzymes in the chain causes conformational changes in the proteins that alter the affinities of some amino acid side-chain gronps for protons. In addition, there is a change in the direction in which these groups face in the membrane. Consequently, oxidation results in an association with a proton on the matrix side of the membrane whereas reduction results in reversal of the direction that the side-chain groups face and an increase in... 

Similarly, this amphiphilic polymer micelle was also used to dismpt the complex between cytochrome c (Cc) and cytochrome c peroxidase (CcP Sandanaraj, Bayraktar et al. 2007). In this case, we found that the polymer modulates the redox properties of the protein upon binding. The polymer binding exposes the heme cofactor of the protein, which is buried in the protein and alters the coordination environment of the metal. The exposure of heme was confirmed by UV-vis, CD spectroscopy, fluorescence spectroscopy, and electrochemical kinetic smdies. The rate constant of electron transfer (fc°) increased by 3 orders of magnimde for the protein-polymer complex compared to protein alone. To establish that the polymer micelle is capable of disrupting the Cc-CcP complex, the polymer micelle was added to the preformed Cc-CcP complex. The observed for this complex was the same as that of the Cc-polymer complex, which confirms that the polymer micelle is indeed capable of disrupting the Cc-CcP complex. 

In natural systems, redox proteins such as cytochrome c (cyt c) function not only to transfer electrons, but to transfer electrons specifically to a particular redox partner, usually another macromolecule. Transfer of electrons between subunits of modified hemoglobins and within complexes of cyt c with cyt b5 and cyt c with cyt c peroxidase (Cep) have therefore been studied extensively (41,42). These studies have revealed the fundamental requirements for the recognition process leading to the formation of the protein-protein complex as well as the thermodynamic features of the electron-transfer reaction itself. This reaction, outlined in equation 6, consists of three fundamental processes recognition to form a complex (Ki), electron transfer within the complex, and dissociation of the redox-altered complex (K2). For the cyt c-Ccp complex, Fe(II) cyt c corresponds to P2red and oxidized Cep corresponds to P °x. 

The disulfide formation depends on the protein conformation that places Cys residues into appropriate proximity and the disulfide redox potential that determines the intrinsic stability of protein disulfide bonds. For catalytic activity, the reduced dithiol form of protein disulfide isomerase is required. Protein disulfide isomerase is a folding catalyst that assists protein folding (Gilbert, 1997). The enzyme increases the rate of the overall folding process of the substrate protein without altering its pathway. 

Obviously there are differences between metal surfaces and metalloporphyrins as well. The electron gas of a metal contains moles of electrons whereas that of a porphyrin is restricted to approximately 25 electrons. This gives a continuous density of states for a metal but discrete bands for a porphyrin. Electron stoichiometry, which is essential in the redox reactions of proteins, is less relevant on metal surfaces, since the Fermi sea is a practically unlimited buffer of electrons. The substitution of an amino-add in the protein may alter the relative affinities for carbon monoxide and dioxygen between two hemoglobins without much, or any, shift of Eq. This is perceived as steric influences or direct impacts on the liganded molecule from the surrounding proteins. Moreover, metal surfaces provide binding sitra of different symmetries, which can be highly selective in certain reactions. Nevertheless, mainly because of the response of... 

The Na/K ATPase has been extensively purified and characterized, and consists of a catalytic a subunit of around 95 kDa and a glycoprotein 0 subunit of approximately 45 kDa (Skou, 1992). The functional transporter exists as a dimer with each monomer consisting of an a and /3 subunit. Hiatt aal. (1984) have su ested that the non-catalytic jS subunit may be involved in the cottect insertion of the a subunit into the lipid bilayer and, therefore, it is conceivable that a modification of the 0 subunit structure may be reflected by changes in the catalytic activity of the a subunit. Therefore, in studies involving the manipulation of tissue glutathione levels, alterations of intracellular redox state may have an effect on substrate binding at an extracellular site on this ion-translocating protein. 

If cellular redox state, determined by the glutathione status of the heart, plays a role in the modulation of ion transporter activity in cardiac tissue, it is important to identify possible mechanisms by which these effects are mediated. Protein S-,thiolation is a process that was originally used to describe the formation of adducts of proteins with low molecular thiols such as glutathione (Miller etal., 1990). In view of the significant alterations of cardiac glutathione status (GSH and GSSG) and ion-transporter activity during oxidant stress, the process of S-thiolation may be responsible for modifications of protein structure and function. 

The nature of the ligand donor atom and the stereochemistry at the metal ion can have a profound effect on the redox potential of redox-active metal ions. The standard redox potentials of Cu2+/Cu+, Fe3+/Fe2+, Mn3+/Mn2+, Co3+/Co2+, can be altered by more than 1.0 V by varying such parameters. A simple example of this effect is provided by the couple Cu2+/Cu+. These two forms of copper have quite different coordination geometries, and ligand environments, which are distorted towards the Cu(I) geometry, will raise the redox potential, as we will see later in the case of the electron transfer protein plastocyanin. 

The ferro-complex CD spectrum shows that reduction of the heme iron alters the heme environment. Redox-induced protein conformation changes could alter the S5unmetry in the heme pocket or produce two binding modes for the reduced complex whose asymmetries nearly cancel each other. Redox-linked conformational changes are especially interesting in view of recent findings of oxido-reductase activity associated with the heme-hemopexin-receptor interaction (89). 

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