43 research outputs found
Copper-sulfenate complex from oxidation of a cavity mutant of Pseudomonas aeruginosa azurin
Metal-sulfenate centers are known to play important roles in biology and yet only limited examples are known due to their instability and high reactivity. Herein we report a copper-sulfenate complex characterized in a protein environment, formed at the active site of a cavity mutant of an electron transfer protein, type 1 blue copper azurin. Reaction of hydrogen peroxide with Cu(I)-M121G azurin resulted in a species with strong visible absorptions at 350 and 452 nm and a relatively low electron paramagnetic resonance gz value of 2.169 in comparison with other normal type 2 copper centers. The presence of a side-on copper-sulfenate species is supported by resonance Raman spectroscopy, electrospray mass spectrometry using isotopically enriched hydrogen peroxide, and density functional theory calculations correlated to the experimental data. In contrast, the reaction with Cu(II)-M121G or Zn(II)-M121G azurin under the same conditions did not result in Cys oxidation or copper-sulfenate formation. Structural and computational studies strongly suggest that the secondary coordination sphere noncovalent interactions are critical in stabilizing this highly reactive species, which can further react with oxygen to form a sulfinate and then a sulfonate species, as demonstrated by mass spectrometry. Engineering the electron transfer protein azurin into an active copper enzyme that forms a copper-sulfenate center and demonstrating the importance of noncovalent secondary sphere interactions in stabilizing it constitute important contributions toward the understanding of metal-sulfenate species in biological systems
Copper-sulfenate complex from oxidation of a cavity mutant of Pseudomonas aeruginosa azurin
Metal-sulfenate centers are known to play important roles in biology and yet only limited examples are known due to their instability and high reactivity. Herein we report a copper-sulfenate complex characterized in a protein environment, formed at the active site of a cavity mutant of an electron transfer protein, type 1 blue copper azurin. Reaction of hydrogen peroxide with Cu(I)-M121G azurin resulted in a species with strong visible absorptions at 350 and 452 nm and a relatively low electron paramagnetic resonance gz value of 2.169 in comparison with other normal type 2 copper centers. The presence of a side-on copper-sulfenate species is supported by resonance Raman spectroscopy, electrospray mass spectrometry using isotopically enriched hydrogen peroxide, and density functional theory calculations correlated to the experimental data. In contrast, the reaction with Cu(II)-M121G or Zn(II)-M121G azurin under the same conditions did not result in Cys oxidation or copper-sulfenate formation. Structural and computational studies strongly suggest that the secondary coordination sphere noncovalent interactions are critical in stabilizing this highly reactive species, which can further react with oxygen to form a sulfinate and then a sulfonate species, as demonstrated by mass spectrometry. Engineering the electron transfer protein azurin into an active copper enzyme that forms a copper-sulfenate center and demonstrating the importance of noncovalent secondary sphere interactions in stabilizing it constitute important contributions toward the understanding of metal-sulfenate species in biological systems
Odorranalectin Is a Small Peptide Lectin with Potential for Drug Delivery and Targeting
BACKGROUND: Lectins are sugar-binding proteins that specifically recognize sugar complexes. Based on the specificity of protein-sugar interactions, different lectins could be used as carrier molecules to target drugs specifically to different cells which express different glycan arrays. In spite of lectin's interesting biological potential for drug targeting and delivery, a potential disadvantage of natural lectins may be large size molecules that results in immunogenicity and toxicity. Smaller peptides which can mimic the function of lectins are promising candidates for drug targeting. PRINCIPAL FINDINGS: Small peptide with lectin-like behavior was screened from amphibian skin secretions and its structure and function were studied by NMR, NMR-titration, SPR and mutant analysis. A lectin-like peptide named odorranalectin was identified from skin secretions of Odorrana grahami. It was composed of 17 aa with a sequence of YASPKCFRYPNGVLACT. L-fucose could specifically inhibit the haemagglutination induced by odorranalectin. (125)I-odorranalectin was stable in mice plasma. In experimental mouse models, odorranalectin was proved to mainly conjugate to liver, spleen and lung after i.v. administration. Odorranalectin showed extremely low toxicity and immunogenicity in mice. The small size and single disulfide bridge of odorranalectin make it easy to manipulate for developing as a drug targeting system. The cyclic peptide of odorranalectin disclosed by solution NMR study adopts a beta-turn conformation stabilized by one intramolecular disulfide bond between Cys6-Cys16 and three hydrogen bonds between Phe7-Ala15, Tyr9-Val13, Tyr9-Gly12. Residues K5, C6, F7, C16 and T17 consist of the binding site of L-fucose on odorranalectin determined by NMR titration and mutant analysis. The structure of odorranalectin in bound form is more stable than in free form. CONCLUSION: These findings identify the smallest lectin so far, and show the application potential of odorranalectin for drug delivery and targeting. It also disclosed a new strategy of amphibian anti-infection
Redox-dependent structural changes in an engineered heme-copper center in myoglobin: Insights into chloride binding to CUB in heme copper oxidases
The effects of chloride on the redox properties of an engineered binuclear heme-copper center in myoglobin (CuBMb) were studied by UV-vis spectroelectrochemistry and EPR spectroscopy. A low-spin heme Fe III-CuI intermediate was observed during the redox titration of CuBMb only in the presence of both CuII and chloride. Upon the first electron transfer to the CuB center, one of the His ligands of CuB center dissociates and coordinates to the heme iron, forming a six-coordinate low-spin ferric heme center and a reduced CUB center. The second electron transfer reduces the ferric heme and causes the release of the coordinated His ligand. Thus, the fully reduced state of the heme-copper center contains a five-coordinate ferrous heme and a reduced CUB center, ready for O2 binding and reduction to water to occur. In the absence of a chloride ion, formation of the low-spin heme species was not observed. These redox reactions are completely reversible. These results indicate that binding of chloride to the CUB center can induce redox-dependent structural changes, and the bound chloride and hydroxide in the heme-copper center may play different roles in the redox-linked enzymatic reactions of heme-copper oxidases, probably because of their different binding affinity to the copper center and the relatively high concentration of chloride under physiological conditions. Β© 2005 American Chemical Society
Radiation-induced defects in apophyllites. II. An O centre and related OO pairs in hydroxylapophyllite
Single-crystal EPR and ENDOR study of an AlO center in prehnite: implications for aluminum-associated oxyradicals in layer silicates
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Coal and Coal Constituent Studies by Advanced EMR Techniques
Advanced electronic magnetic resonance (EMR) methods are used to examine properties of coals, chars, and molecular species related to constituents of coal. We have achieved substantial progress on upgrading the high field (HF) EMR (W-band, 95 GHz) spectrometers that are especially advantageous for such studies. Particularly, we have built a new second W-band instrument (Mark II) in addition to our Mark I. Briefly, Mark II features: (i) an Oxford custom-built 7 T superconducting magnet which is scannable from 0 to 7 T at up to 0.5 T/min; (ii) water-cooled coaxial solenoid with up to Β±550 G scan under digital (15 bits resolution) computer control; (iii) custom-engineered precision feed-back circuit, which is used to drive this solenoid, is based on an Ultrastab 860R sensor that has linearity better than 5 ppm and resolution of 0.05 ppm; (iv) an Oxford CF 1200 cryostat for variable temperature studies from 1.8 to 340 K. During this grant period we have completed several key upgrades of both Mark I and II, particularly microwave bridge, W-band probehead, and computer interfaces. We utilize these improved instruments for HF EMR studies of spin-spin interaction and existence of different paramagnetic species in carbonaceous solids