Restrained least squares refinement of native (calcium) and cadmium-substituted carp parvalbumin using X-ray crystallographic data at 1.6-A resolution.

Carp parvalbumin coordinates calcium through one carbonyl oxygen atom and the oxygen-containing side chains of 5 amino acid residues, or 4 residues and a water molecule, in a helix-loop-helix structural motif. Other calcium-binding proteins, including calmodulin and troponin C, also possess this unique calcium-binding design, which is designated EF-hand or calmodulin fold. Parvalbumin has two such sites, labeled CD and EF. Each of the calcium-binding sites of refined structures of proteins belonging to this group has a 7-oxygen coordination sphere except those of the structure of parvalbumin as it was reported in 1975. This structure had been refined at 1.9 A using difference Fourier techniques on film data. The CD site appeared to be 6-coordinate and the EF site 8-coordinate. Results of NMR experiments using 113Cd-substituted parvalbumin, however, indicate that the sites are similar to one another with coordination number greater than 6. To resolve the inconsistency between crystallographic and NMR results, 1.6 A area detector data was collected for native and cadmium-substituted parvalbumin; the structures have been refined to R factors of 18.7% and 16.4%, respectively, with acceptable geometry and low errors in atomic coordinates. Differences between the parvalbumin structure described in 1975 and the present structure are addressed, including the discovery of 7-coordination for both the CD and EF sites.

Superoxide dismutase-like activities of copper(II) complexes tested in serum.

The superoxide dismutase-like activities of a series of coordination complexes of copper were evaluated and compared to the activities of bovine erythrocyte superoxide dismutase (superoxide: superoxide oxidoreductase, EC 1.15.1.1) in serum using the nitroblue tetrazolium chloride (NBT)-reduction assay and electron paramagnetic resonance (EPR) spectroscopy. A 40% inhibition was observed for the initial rate of the NBT reduction by superoxide dismutase in serum, but more than 40% inhibition was achieved with CuSO4, Cu(II)-dimethylglyoxime, Cu(II)-3,8-dimethyl-4,7-diazadeca-3,7-dienediamide, Cu2[N,N'-(2-(O-hydroxy-benzhydrylidene)amino)ethyl]2-1,2-ethane dia mine), Cu(II)-(diisopropylsalicylate)2, Cu(II)-(p-bromo-benzoate)2, Cu(II)-(nicotinate)2 and Cu(II)-(1,2-diamino-2-methylpropane)2. The electron paramagnetic resonance technique of spin trapping was used to detect the formation of superoxide (O2-.) and other free radicals in the xanthine-xanthine oxidase system under a variety of conditions. Addition of the spin trapping agent 5,5-dimethylpyrroline 1-oxide (DMPO) to the xanthine-xanthine oxidase system in fetal bovine serum produced the O2-.-spin adduct of DMPO (herein referred to as superoxide spin adduct, DMPO-OOH) as the well known short-lived nitroxyl whose characteristic EPR spectrum was recorded before its rapid decay to undetectable levels. The hydroxyl radical (HO.) adduct of the spin trap DMPO (herein referred to as DMPO-OH) was detected to a very small extent. When CuSO4, or the test complexes of copper, were added to the xanthine-xanthine oxidase system in serum containing the spin trap, the yield of DMPO-OOH was negligible. In addition to their superoxide dismutase-like activity, CuSO4 and the copper complexes also behaved as Fenton-type catalysts as seen by the accumulation of varying amounts of the hydroxyl spin adduct DMPO-OH. Both the Fenton-type catalysis and the superoxide dismutase-like action of these compounds were lost when a chelator such as EDTA was included in the xanthine-xanthine oxidase incubation mixture. Addition of superoxide dismutase instead of the copper compounds to this enzyme system abolished the formation of superoxide adduct DMPO-OOH, and no hydroxyl adduct DMPO-OH was detected. This effect of superoxide dismutase remained unaltered by EDTA.

Structure of human hemoglobin C: a disease with intraerythrocytic crystals.

Crystals of human cyanomethemoglobin C (beta 6A3 glu leads to Lys) crystallized in the orthorhombic space group P212121, A = 158(1), B = 65.5(4), C = 54.9(5) A with Z =4. Single crystal electron micrographs show filaments parallel to the b direction. The molecules are unusually densely packed compared to other hemoglobin crystals, and this may be related to the ease of intraerythrocytic crystallization.

Crystallography and oriented single crystal electron microscopy of hemoglobin deer II, a hemoglobin that exhibits matchstick-shaped erythrocytes.

Deer hemoglobin beta chain type II has been crystallized and preliminary diffraction data and oriented single crystal transmission electron micrographs have been obtained. The crystals are monoclinic P21 with Z = 4. The electron micrographs show a herringbonelike structure in the ab plane with open rectangular solvent channels and a fiber-like arrangement of molecules perpendicular to this plane.