Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A heme protein, hCP, from the extreme halophile, Haloarcula marismortui, showing both peroxidatic and catalatic activity has been purified and characterized as a catalase-peroxidase. Catalatic activity is enhanced by molar concentrations of NaCl or (NH4)2SO4, while peroxidase activity decreases with increasing salt concentration. Optimal pH values are 6.0 for peroxidatic activity assayed in absence of NaCl and 7.5 for catalatic activity assayed in molar concentrations of NaCl. The two activities present saturation behaviour with increasing H2O2 concentration with apparent Km values of 0.5 and 2.5 mM for the peroxidatic and catalatic activities, respectively. A molecular mass of 81,292 +/- 9 Da was measured for the polypeptide by mass spectroscopy. One heme group (protoporphyrin IX with an iron atom in the ferric state) is associated with one molecule of hCP. Its amino-acid composition shows hCP to contain a high proportion of acidic residues. The EPR spectrum of the NO-compound of reduced (ferrous) hCP strongly suggests that the proximal ligand of the heme is the imidazole group of a histidine residue.
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PMID:Purification and properties of a halophilic catalase-peroxidase from Haloarcula marismortui. 794 69

A new ferredoxin has been purified from the photosynthetic bacterium Rhodobacter capsulatus. It is the sixth ferredoxin to be isolated from this bacterium and it was called FdVI. Its primary structure was established based on amino acid sequence analysis of the protein and of peptides derived from it. It is composed of 106 residues including five cysteines. The calculated mass of the polypeptide is 11,402.6 Da which matches the experimental value determined by electrospray mass spectrometry. Amino acid sequence comparison revealed that ferredoxin VI (FdVI) is strikingly similar to a ferredoxin from Caulobacter crescentus and to the putidaredoxin from Pseudomonas putida. FdVI exhibited an ultraviolet-visible absorption spectrum typical for a [2Fe-2S] ferredoxin. EPR spectroscopy of the reduced protein showed a nearly axial signal similar to that of mitochondrial and P. putida ferredoxins. FdVI is biosynthesized in cells growing anaerobically under either nitrogen-sufficient or nitrogen-deficient conditions. Although the function of FdVI is unknown, its structural resemblance to [2Fe-2S] ferredoxins known to transfer electrons to oxygenases such as P-450 cytochromes, suggests that FdVI may have a similar role in R. capsulatus.
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PMID:Purification of a sixth ferredoxin from Rhodobacter capsulatus. Primary structure and biochemical properties. 802 3

Jujuboside A (JuA), an effective component of sanzaoren, a Chinese herbal medicine, is a noncompetitive inhibitor of calmodulin (CaM). The interaction of JuA with CaM has been investigated with 1H NMR and spin-labeled EPR spectroscopies. The 1H NMR experiments showed that JuA has two kinds of binding sites on CaM: one locates in the N-terminus, the other locates in the C-terminal region of the polypeptide chain. The EPR studies on 3-maleimido-2,2,5,5-tetramethyl-1-pyrrolidinyloxyl labeled CaM (spin-labeled CaM) revealed that each CaM molecule can bind at least two JuA molecules. Binding of JuA affects the environments of some lysine residues (most likely Lys-74 and Lys-94), suggesting that JuA binds to CaM through hydrophobic interaction.
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PMID:1H NMR and spin-labeled EPR studies on the interaction of calmodulin with jujuboside A. 803 6

Overexpression of foreign proteins in Escherichia coli is usually tested by sodium dodecyl sulfate-polyacrylamide gel electrophoresis or Western blot analysis. However, when metalloproteins of flavoenzymes are expressed, the correct assembly of the polypeptide chain to the cofactor cannot be verified using these methods. We have used EPR spectroscopy and Mancini tests to monitor the expression of holoflavodoxin from the cyanobacteria Anabaena in E. coli. Flavodoxin from Anabaena sp PCC 7119 was cloned in the plasmid pTrc 99b after the trc promotor, which is activated in the presence of isopropyl-beta-D-thiogalactoside. Two hours after induction, most of the recombinant apoflavodoxin is already synthesized. However, only 65% of this protein is assembled to flavin mononucleotide (FMN). Harvesting of the cells to obtain all the flavodoxin in the holo form must be carried out 10 h after induction. Addition of FMN to the culture media increases the synthesis of holoflavodoxin by about 17%. The presence of flavin adenine dinucleotide or riboflavin in the culture inhibits the accumulation of semiquinone flavodoxin in the cells.
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PMID:Electron paramagnetic resonance as a tool for monitoring overexpression in Escherichia coli of fully functional flavodoxin. 807 77

We have cloned the gene encoding Desulfovibrio gigas ferredoxin using a photodigoxigenin-labelled probe synthesized with the polymerase chain reaction. The DNA sequence of the gene predicts a polypeptide of 58 residues after removal of the initial formyl methionine (polypeptide M(r) = 6,276). The ferredoxin gene was expressed in aerobically grown E. coli behind the lac promoter of pUC18 resulting in a high level of ferredoxin expression which comprises about 10% of the total cell protein. EPR analysis of recombinant ferredoxin revealed the presence of a [3Fe-4S] cluster which is characteristic of native D. gigas ferredoxin II.
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PMID:Cloning, sequencing and overexpression of the Desulfovibrio gigas ferredoxin gene in E. coli. 808 3

The reaction center of photosystem II of oxygenic photosynthesis contains two redox-active tyrosines called Z and D, each of which can act as an electron donor to the oxidized primary electron donor, P680+. These tyrosines are located in homologous positions on the third transmembrane alpha-helix of each of the two homologous polypeptides, D1 and D2, that comprise the reaction center. Tyrosine D of polypeptide D2 has been proposed, upon oxidation, to give up its phenolic proton to a nearby basic amino acid residue, forming a neutral radical. Modeling studies have pointed to His190 (spinach numbering) as a likely candidate for this basic residue. As a test of this hypothesis, we have constructed three site-directed mutations in the D2 polypeptide of the cyanobacterium Synechocystis sp. PCC6803. His189 (the Synechocystis homologue of His190 of spinach) has been replaced by glutamine, aspartate, or leucine. Instead of the normal D. EPR signal (g = 2.0046; line width 16-19 G), PSII core complexes isolated from these three mutants show an altered dark-stable EPR signal with a narrowed line width (11-13 G), and g values of 2.0046, 2.0043, and 2.0042 for the His189Gln, His189Asp, and His189Leu mutants, respectively. Despite the reduced line width, these EPR signals show g values and microwave-power saturation properties similar to the normal D. signal. Furthermore, specific deuteration in one of those mutants at the 3 and 5 positions of the phenol ring of the photosystem II reaction center tyrosines results in a loss of hyperfine structure of the EPR signal, proving that the signal indeed arises from tyrosine.2+ This observation provides support for a model in which an imidazole nitrogen of His189 accepts the phenolic proton of Tyr160 upon oxidation of D, forming a back hydrogen bond to the phenolic oxygen of the neutral tyrosyl radical.
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PMID:Spectroscopic evidence from site-directed mutants of Synechocystis PCC6803 in favor of a close interaction between histidine 189 and redox-active tyrosine 160, both of polypeptide D2 of the photosystem II reaction center. 825 9

Desulfoferrodoxin from Desulfovibrio vulgaris, strain Hildenborough, is a homodimer of 28 kDa; it contains two Fe atoms per 14.0 kDa subunit. The N-terminal amino-acid sequence is homogeneous and corresponds to the previously described Rho gene, which encodes a highly charged 14 kDa polypeptide without a leader sequence. Although one of the two iron centers, FeA, has previously been described as a 'strained rubredoxin-like' site, EPR of the ferric form proves very similar to that of the pentagonal bipyramidally coordinated iron in ferric complexes of DTPA, diethylenetriaminepentaacetic acid: both systems have spin S = 5/2 and rhombicity E/D = 0.08. Unlike the Fe site in rubredoxin the FeA site in desulfoferrodoxin has a pH dependent midpoint potential with pKox = 9.2 and pKred = 5.3. Upon reduction (Em,7.5 = +2 mV) FeA exhibits an unusually sharp S = 2 resonance in parallel-mode EPR. The second iron, FeB, has S = 5/2 and E/D = 0.33; upon reduction (Em,7.5 = +90 mV) FeB turns EPR-silent.
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PMID:On the two iron centers of desulfoferrodoxin. 826 95

Deletion of the nifW gene in Azotobacter vinelandii yields a strain (DJ224) that grows poorly under N2 fixing conditions (Jacobson, M. R., Cash, V. L., Weiss, M. C., Laird, N. F., Newton, W. E., and Dean, D. R. (1989) Mol. & Gen. Genet. 219, 49-57). Here we report the purification of nitrogenase from DJ224. The purified Fe protein was indistinguishable from wild-type. The MoFe protein was indistinguishable from the wild-type MoFe protein by the criteria of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, native gel electrophoresis, two-dimensional gel electrophoresis, metal analysis, UV/visible and EPR spectroscopies. It was different by the criteria of CD spectroscopy and specific activities. At a 5:1 molar ratio of Fe protein to MoFe protein, H2 evolution under argon was identical to wild-type, C2H2 reduction was inhibited by 27%, N2 reduction was inhibited by 38%, and CO inhibited H2 evolution by 17%. The above data show that the nifW gene product is not required for: 1) detectable alteration of the polypeptide; 2) the synthesis of the metal portion of FeMo cofactor; or 3) FeMo cofactor insertion. The MoFe protein synthesized in the absence of NifW appears to have an alteration near the FeMo cofactor site, possibly at homocitrate, which causes differential inhibition of different substrates.
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PMID:Purification and characterization of nitrogenase from a delta nifW strain of Azotobacter vinelandii. 830 84

Photosystem II contains two redox-active tyrosines, D and Z. To understand the function of the dark stable tyrosine radical, D+, we have characterized two site-directed mutations at the D tyrosine residue in the transformable cyanobacterium, Synechocystis sp. PCC 6803, through the use of purified photosystem II particles (Noren, G. H., Boerner, R. J., and Barry, B. A. (1991) Biochemistry 30, 3943-3950). In manganese-depleted mutant particles, a light-induced EPR signal is observed. This signal contains a stable component, due to a chlorophyll radical, and an unstable component. The lineshape of the unstable, oxidized component, which we call M+, is obtained by subtraction; it has a lineshape different from tyrosine Z+/D+ and a g value of 2.004. Up to one M+ spin per reaction center can be photooxidized. The characteristic light-induced EPR signal ascribed to Z+ is not detected; under the same conditions, Z+ is detected in control preparations. The M+ radical lineshape is similar to the light-induced photosystem II radical identified in a site-directed mutant in the D1 polypeptide (YF161D1) (Noren, G. H., and Barry, B. A. (1992) Biochemistry 31, 3335-3342). Optical measurements on manganese-depleted photosystem II particles from control and D2 mutant preparations show that charge recombination kinetics between Q-A and an oxidized redox-active component are similar, to within a factor of two, in all three preparations. We conclude that lack of the stable tyrosine D+ alters the structure or redox properties of tyrosine Z in manganese-depleted preparations.
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PMID:Removal of stable tyrosine radical D+ affects the structure or redox properties of tyrosine Z in manganese-depleted photosystem II particles from Synechocystis 6803. 838 May 79

Anaerobically grown Escherichia coli contain an oxygen-sensitive ribonucleotide reductase. The enzyme requires anaerobic activation by two E. coli fractions with S-adenosylmethionine, NADPH, dithiothreitol, and KCl. We now find that photochemically reduced deazaflavin can substitute for these two fractions and NADPH. The reductase contained roughly equimolar amounts of iron and sulfide, suggesting the presence of an Fe-S complex. The cluster is characterized by a charge transfer band at 420 nm and a low temperature EPR signal centered at g = 2.01 that is difficult to saturate at 14 K, suggested to be a (3Fe-4S)+ cluster. In five different preparations of essentially protein-pure reductase containing widely different amounts of iron, the catalytic activity correlated well with the iron content. The iron signal disappeared during reductive anaerobic activation, with the appearance of a new EPR signal at g = 2.0033 showing a temperature behavior and microwave power saturability consistent with an organic free radical. The signal disappeared after exposure of the activated enzyme to air. We suggest that activation involves generation of a specific amino acid free radical that is dependent on the reduced Fe-S cluster and S-adenosylmethionine. From other work it appears likely that the free radical is localized on glycine 681 of the polypeptide chain.
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PMID:An iron-sulfur center and a free radical in the active anaerobic ribonucleotide reductase of Escherichia coli. 838 2


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