Some bacterial species are able to utilize extracellular mineral forms of iron and manganese as respiratory electron acceptors. In Shewanella oneidensis this involves decaheme cytochromes that are located on the bacterial cell surface at the termini of transouter-membrane electron transfer conduits. The cell surface cytochromes can potentially play multiple roles in mediating electron transfer directly to insoluble electron sinks, catalyzing electron exchange with flavin electron shuttles or participating in extracellular intercytochrome electron exchange along "nanowire" appendages. We present a 3.2-Å crystal structure of one of these decaheme cytochromes, MtrF, that allows the spatial organization of the 10 hemes to be visualized for the first time. The hemes are organized across four domains in a unique crossed conformation, in which a staggered 65-Å octaheme chain transects the length of the protein and is bisected by a planar 45-Å tetraheme chain that connects two extended Greek key split β-barrel domains. The structure provides molecular insight into how reduction of insoluble substrate (e.g., minerals), soluble substrates (e.g., flavins), and cytochrome redox partners might be possible in tandem at different termini of a trifurcated electron transport chain on the cell surface.c-type cytochromes | iron respiration | MtrC | multiheme

Ferritins are a superfamily of iron oxidation, storage and mineralization proteins found throughout the animal, plant, and microbial kingdoms. The majority of ferritins consist of 24 subunits that individually fold into 4-α-helix bundles and assemble in a highly symmetric manner to form an approximately spherical protein coat around a central cavity into which an iron-containing mineral can be formed. Channels through the coat at inter-subunit contact points facilitate passage of iron ions to and from the central cavity, and intrasubunit catalytic sites, called ferroxidase centers, drive Fe2+ oxidation and O2 reduction. Though the different members of the superfamily share a common structure, there is often little amino acid sequence identity between them. Even where there is a high degree of sequence identity between two ferritins there can be major differences in how the proteins handle iron. In this review we describe some of the important structural features of ferritins and their mineralized iron cores, consider how iron might be released from ferritins, and examine in detail how three selected ferritins oxidise Fe2+ to explore the mechanistic variations that exist amongst ferritins. We suggest that the mechanistic differences reflect differing evolutionary pressures on amino acid sequences, and that these differing pressures are a consequence of different primary functions for different ferritins.

a b s t r a c tMagnetic circular dichroism (MCD) spectra, at ultraviolet-visible or near-infrared wavelengths (185-2000 nm), contain the same transitions observed in conventional absorbance spectroscopy, but their bisignate nature and more stringent selection rules provide greatly enhanced resolution. Thus, they have proved to be invaluable in the study of many transition metal-containing proteins. For mainly technical reasons, MCD has been limited almost exclusively to the measurement of static samples. But the ability to employ the resolving power of MCD to follow changes at transition metal sites would be a potentially significant advance. We describe here the development of a cuvette holder that allows reagent injection and sample mixing within the 50-mm-diameter ambient temperature bore of an energized superconducting solenoid. This has allowed us, for the first time, to monitor time-resolved MCD resulting from in situ chemical manipulation of a metalloprotein sample. Furthermore, we report the parallel development of an electrochemical cell using a three-electrode configuration with physically separated working and counter electrodes, allowing true potentiometric titration to be performed within the bore of the MCD solenoid.Ó 2011 Elsevier Inc. All rights reserved.Magnetic circular dichroism (MCD) 1 spectroscopy, at ultraviolet-visible and near-infrared wavelengths (185-2000 nm), has proved to be invaluable in the study of metalloproteins containing cofactors such as heme [1][2][3], non-heme iron [4], iron-sulfur clusters [5][6][7][8], cobalt [9,10], nickel [11][12][13], and copper [14,15]. The technique measures the apparent circular dichroism (CD) induced by a magnetic field [16]. Despite similarities in the instrumentation used, the observation of MCD is not dependent on the chirality of the protein; the method is equally applicable to racemic model complexes [17]. The magnetic field will always induce signals across wavelengths at which the substance absorbs. Thus, MCD spectra contain the same electronic transitions observed in conventional absorbance spectroscopy, but the bisignate nature of the spectrum provides enhanced resolution and greater detail. This spectral detail offers an unmatched fingerprinting capability that can, for example, identify the spin and oxidation states of heme groups [1] and distinguish among the variety of iron-sulfur centers found in biological molecules [18][19][20].Metalloprotein MCD can be measured at ambient or cryogenic temperatures. The latter requires adulteration with glassing agents [16] but has generally been preferred because MCD intensity from paramagnetic centers increases dramatically at low temperature. Thus, most metalloprotein MCD reported has been measured in glasses at temperatures of approximately 4.2 K. Hemoproteins represent the significant exception, giving rise to appreciable MCD at high temperatures [1]. Thus, ambient temperature MCD is used to diagnose spin state, oxidation state, and (in the case of low-spin Fe(III) hemes) axial ligation [21].As ...

Significant progress has been made in recent years toward understanding the processes by which an iron mineral is deposited within members of the ferritin family of 24mer iron storage proteins, enabled by high-resolution structures together with spectroscopic and kinetic studies. These suggest common characteristics that are shared between ferritins, namely, a highly symmetric arrangement of subunits that provides a protein coat around a central cavity in which the mineral is formed, channels through the coat that facilitate ingress and egress of ions, and catalytic sites, called ferroxidase centers, that drive Fe(2+) oxidation. They also reveal significant variations in both structure and mechanism amongst ferritins. Here, we describe three general types of structurally distinct ferroxidase center and the mechanisms of mineralization that they are associated with. The highlighted variation leads us to conclude that there is no universal mechanism by which ferritins function, but instead there exists several distinct mechanisms of ferritin iron mineralization.