X-ray, ESR, and quantum mechanics studies unravel a spin well in the cofactor-less urate oxidase.

Urate oxidase (EC 1.7.3.

Near-atomic resolution structures of urate oxidase complexed with its substrate and analogues: the protonation state of the ligand.

Urate oxidase (uricase; EC 1.7.3.

Structural analysis of urate oxidase in complex with its natural substrate inhibited by cyanide: mechanistic implications.

Background: Urate oxidase (EC 1.7.3.

Oxygen pressurized X-ray crystallography: probing the dioxygen binding site in cofactorless urate oxidase and implications for its catalytic mechanism.

The localization of dioxygen sites in oxygen-binding proteins is a nontrivial experimental task and is often suggested through indirect methods such as using xenon or halide anions as oxygen probes. In this study, a straightforward method based on x-ray crystallography under high pressure of pure oxygen has been developed. An application is given on urate oxidase (UOX), a cofactorless enzyme that catalyzes the oxidation of uric acid to 5-hydroxyisourate in the presence of dioxygen.

Recapture of [S]-allantoin, the product of the two-step degradation of uric acid, by urate oxidase.

Urate oxidase from Aspergillus flavus catalyzes the degradation of uric acid to [S]-allantoin through 5-hydroxyisourate as a metastable intermediate. The second degradation step is thought either catalyzed by another specific enzyme, or spontaneous. The structure of the enzyme was known at high resolution by X-ray diffraction of I222 crystals complexed with a purine-type inhibitor (8-azaxanthin).

Three-dimensional structure of nucleoside diphosphate kinase.

Three-dimensional structures are known from X-ray studies of the nucleoside diphosphate (NDP) kinase of many organisms from bacteria to human. All NDP kinases have subunits of about 150 residues with a very similar fold based on the alphabeta sandwich or ferredoxin fold. This fold is found in many nucleotide or polynucleotide-binding proteins with no sequence relationship to NDP kinase.

Kinetic properties of dihydrofolate reductase from wild-type and mutant Plasmodium vivax expressed in Escherichia coli.

Antifolate drugs inhibit malarial dihydrofolate reductase (DHFR). In Plasmodium falciparum, antifolate resistance has been associated with point mutations in the gene encoding DHFR. Recently, mutations at homologous positions have been observed in the P.

The human nm23-H4 gene product is a mitochondrial nucleoside diphosphate kinase.

We demonstrate here the catalytic activity and subcellular localization of the Nm23-H4 protein, product of nm23-H4, a new member of the human nm23/nucleoside diphosphate (NDP) kinase gene family (Milon, L., Rousseau-Merck, M., Munier, A.

Redox signalling in the chloroplast: structure of oxidized pea fructose-1,6-bisphosphate phosphatase.

Sunlight provides the energy source for the assimilation of carbon dioxide by photosynthesis, but it also provides regulatory signals that switch on specific sets of enzymes involved in the alternation of light and dark metabolisms in chloroplasts. Capture of photons by chlorophyll pigments triggers redox cascades that ultimately activate target enzymes via the reduction of regulatory disulfide bridges by thioredoxins. Here we report the structure of the oxidized, low-activity form of chloroplastic fructose-1, 6-bisphosphate phosphatase (FBPase), one of the four enzymes of the Calvin cycle whose activity is redox-regulated by light.

Mechanism of phosphate transfer by nucleoside diphosphate kinase: X-ray structures of the phosphohistidine intermediate of the enzymes from Drosophila and Dictyostelium.

Nucleoside diphosphate kinase (NDP kinase) has a ping-pong mechanism with a phosphohistidine intermediate. Crystals of the enzymes from Dictyostelium discoideum and from Drosophila melanogaster were treated with phosphoramidate, and their X-ray structures were determined at 2.1 and 2.

A simple synthesis of 11,19-oxidosteroids.

The photochemical hypoiodination of cortisol acetonide gave a mixture of 18-iodocortisol acetonide and of the 11 beta,19-oxidoderivative. The proportion of the two products was slightly modified by the reaction temperature. Deprotection of the acetonide group of the 11 beta,19-oxidoderivative gave 11 beta,19-oxido-17 alpha,21-dihydroxy-4-pregnen-3,20-dione which led to the formation of 11 beta,19-oxido-4-androsten-3,17-dione upon treatment with sodium bismutate.

Crystal structure of the Awd nucleotide diphosphate kinase from Drosophila.

Background: Nucleotide diphosphate kinase (NDP kinase) is a phosphate transfer enzyme involved in cell regulation and in animal development. Drosophila NDP kinase is the product of the abnormal wing disc (awd) developmental gene, a point mutation in which can produce the killer of prune (K-pn) conditional lethal phenotype. The highly homologous mammalian genes control metastasis and a human NDP kinase acts as a transcription factor.

Derivation by statistical methods of phase information from multiple-wavelength anomalous diffraction data. Basic questions, 'best' electron-density map, implementation and tests.

A probability distribution of the structure factor is established from the analysis of the effects of errors involved in the multiple-wavelength anomalous diffraction (MAD) method. This probability distribution, derived from those of the intensities, is two-dimensional for acentric reflections and uni-dimensional for centric reflections. It permits, using the centroid of the distribution, the calculation of the modulus and the phase of the 'best' structure factor.

Protein engineering of xylose (glucose) isomerase from Actinoplanes missouriensis. 3. Changing metal specificity and the pH profile by site-directed mutagenesis.

Aldose-ketose isomerization by xylose isomerase requires bivalent cations such as Mg2+, Mn2+, or Co2+. The active site of the enzyme from Actinoplanes missouriensis contains two metal ions that are involved in substrate binding and in catalyzing a hydride shift between the C1 and C2 substrate atoms. Glu 186 is a conserved residue located near the active site but not in contact with the substrate and not with a metal ligand.

Protein engineering of xylose (glucose) isomerase from Actinoplanes missouriensis. 1. Crystallography and site-directed mutagenesis of metal binding sites.

The structure and function of the xylose (glucose) isomerase from Actinoplanes missouriensis have been analyzed by X-ray crystallography and site-directed mutagenesis after cloning and overexpression in Escherichia coli. The crystal structure of wild-type enzyme has been refined to an R factor of 15.2% against diffraction data to 2.

Arginine residues as stabilizing elements in proteins.

Site-specific substitutions of arginine for lysine in the thermostable D-xylose isomerase (XI) from Actinoplanes missouriensis are shown to impart significant heat stability enhancement in the presence of sugar substrates most probably by interfering with nonenzymatic glycation. The same substitutions are also found to increase heat stability in the absence of any sugar derivatives, where a mechanism based on prevention of glycation can no longer be invoked. This rather conservative substitution is moreover shown to improve thermostability in two other structurally unrelated proteins, human copper, zinc-superoxide dismutase (CuZnSOD) and D-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Bacillus subtilis.

Crystal structure study of Opsanus tau parvalbumin by multiwavelength anomalous diffraction.

The crystal structure of a small calcium-binding protein, the parvalbumin IIIf from Opsanus tau in which Tb was substituted for Ca, has been analysed by multiwavelength anomalous diffraction. Data at a resolution of 2.3 A were collected at three wavelengths near the L3 absorption edge of Tb (1.