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Abstract: Detailed structural models of di-cluster seven-iron ferredoxins constitute a valuable resource for folding and stability studies relating the metal cofactors’ role in protein stability. The here reported, hemihedric twinned crystal structure at 2.0Å resolution from Acidianus ambivalens ferredoxin, shows an integral 103 residues, physiologically relevant native form composed by a N-terminal extension comprising a His/Asp Zn2+ site and the ferredoxin (βαβ)2 core, which harbours intact clusters I and II, a [3Fe–4S]1+/0 and a [4Fe–4S]2+/1+ centres. This is in contrast with the previously available ferredoxin structure from Sulfolofus tokodai, which was obtained from an artificial oxidative conversion with two [3Fe–4S]1+/0 centres and poor definition around cluster II. [Copyright &y& Elsevier]

RuvBL1, an evolutionary highly conserved protein related to the AAA+ family of ATPases, has been crystallized using the hanging-drop vapour-diffusion method at 293 K. The crystals are hexagonal and belong to space group P6, with unit-cell parameters a = b = 207.1, c = 60.7 Å and three molecules in the asymmetric unit. [ABSTRACT FROM AUTHOR]

Crystals of the title protein have been produced and preliminary structural analysis has been carried out. The crystals belong to the orthorhombic space group C2221, with unit-cell parameters a = 258.1, b = 340.1, c = 266.5 Å . The protein forms a 24-mer of 20 kDa subunits, which assemble with 432 noncrystallographic symmetry. A total of 36 monomers are found in the asymmetric unit, corresponding to one and a half 24-mers. [ABSTRACT FROM AUTHOR]

The structure of dimeric cytochrome c[sub 3] from the sulfate reducing bacterium Desulfovibrio gigas, diDg, obtained by ab initio methods was further refined to 1.2 Å resolution, giving final reliability factors of R[sub free] = 14.8% and R = 12.4%. This cytochrome is a dimer of tetraheme cytochrome c[sub 3] molecules covalently linked by two solvent-accessible disulfide bridges, a characteristic unique to members of the cytochrome c[sub 3] superfamily. Anisotropic analysis using the semi-rigid TLS method shows different behaviour for analogous loops in each monomer arising from their different packing environments. A detailed sequence and structural comparison with all other known cytochrome c[sub 3] domains in single- and multi-domain cytochromes c[sub 3] shows the presence of structurally conserved regions in this family, despite the high variability of the aminoacid sequence. An internal water molecule is conserved in a common structural arrangement in all c[sub 3] tetraheme domains, indicating a probable electron-transfer pathway between hemes I and II. Unique features of diDg are an internal methionine residue close to heme I and to one of the axial ligands of heme III, where all other structures of the cytochrome c[sub 3] superfamily have a phenylalanine, and a rather unusual CXXXCH heme-binding motif only found so far in this cytochrome. [ABSTRACT FROM AUTHOR]

Cytochrome c' from the obligate methylotroph Methylophilus methylotrophus is a 15 kDa monohaem protein which has a c-type haem covalently linked to the protein chain. Two histidine residues are the axial ligands of the Fe atom in the oxidized form. This cytochrome is one of the few known haem proteins which undergoes a change of spin state of the Fe atom upon reduction, with the detachment of an axial histidine ligand. Initial crystallization conditions involved the utilization of cadmium chloride as an additive and resulted in highly mosaic crystals with poor diffraction properties. Optimization of the crystallization conditions was achieved by slowing the nucleation process utilizing agarose gels and viscous additives such as PEG, ethylene glycol and glycerol. Addition of glycerol to the crystallization buffer produced crystals suitable for X-ray diffraction, with a reduced solvent content and mosaicity, which diffracted to a maximum resolution of 1.19 Å using synchrotron radiation. The crystals obtained under these conditions were employed for structure solution using the multiwavelength anomalous dispersion method at the Fe K edge. [ABSTRACT FROM AUTHOR]

The facultative sulfate/nitrate-reducing bacterium Desulfovibrio desulfuricans ATCC 27774 harbours a split-Soret cytochrome c. This cytochrome is a homodimeric protein, having two bis-histidinyl c -type haems per monomer. It has an unique architecture at the haem domain: each haem has one of the coordinating histidines provided by the other monomer, and in each monomer the haems are parallel to each other, almost in van der Waals contact. This work reports the cloning and sequencing of the gene encoding for this cytochrome and shows, by transcriptional analysis, that it is more expressed in nitrate-grown cells than in sulfate-grown ones. In addition, the gene-deduced amino acid sequence revealed two new cysteine residues that could be involved in the binding of a non-haem iron centre. Indeed, the presence of a novel type of an iron-sulfur centre (possibly of the [2Fe-2S] type) was demonstrated by EPR spectroscopy, and putative models for its localization and structure in the cytochrome molecule are proposed on the basis of the so-far-known 3D crystallographic structure of the aerobically purified split-Soret cytochrome, which lacks this centre. [ABSTRACT FROM AUTHOR]

Assesses the crystallization and X-ray characterization of laccase CotA from Bacillus subtilis. Increase thermostability of the laccase; Synthesis of B. subtilis; Laccase activity of CotA.