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M Bruschi

Publications and source records attributed to M Bruschi.

At least 91 records · Page 5Linked to original sources
PubMed1987

Thermodynamic parameters of cytochrome c3-ferredoxin complex formation.

The complex formation between cytochrome c3 and ferredoxin I from Desulfovibrio desulfuricans Norway was studied by microcalorimetric and pH-stat titration measurements. The stoichiometry of the complex was found to be one molecule of cytochrome c3 per monomer of ferredoxin I. The association constant determined at T = 283 K in tris(hydroxymethyl)aminomethane hydrochloride (Tris-HCl) buffer, 10(-2) M and pH 7.7, was KA = 1.3 X 10(6) M-1. Though the enthalpy (delta H = 19 +/- 1 kJ.mol-1) and the entropy (delta S = 183 J.K-1.mol-1) were positive and consistent with a hydrophobic process involved in the interaction, the analysis of ionic strength dependence exhibited an important electrostatic effect on the association. The use of both Tris-HCl and phosphate buffers during microcalorimetric experiments showed proton release at pH 6.6. The pH-stat study of proton release indicated that one of the charged groups involved in the interacting site underwent a pK shift from 7.35 to 6.05.

Calorimetry↗
PubMed1987

Rubredoxin from Desulfovibrio gigas. A molecular model of the oxidized form at 1.4 A resolution.

The crystal structure of rubredoxin from the sulfate-reducing bacterium Desulfovibrio gigas has been determined at 1.4 A resolution (1 A = 0.1 nm) by X-ray diffraction methods; starting with a model of the isostructural rubredoxin from Desulfovibrio vulgaris. Refinement of the molecular model has been carried out by restrained least-squares techniques and Fourier series calculations. The present model includes a formyl at the N-terminal end and 121 possible sites for solvent molecules with full or partial occupancy, which corresponds to the modeling of nearly all the solvent medium. The crystallographic R factor against the data with 10 A greater than d greater than 1.4 A with F greater than 2 sig(F), is 0.136; and R = 0.140 when all the data are considered. The estimated average root-mean-square (r.m.s.) error on the positional parameters is about 0.12 A. The overall structural features of this molecule are close to those of the two highly refined rubredoxins from Clostridium pasteurianum and D. vulgaris. Superposition of these two molecules on the rubredoxin from D. gigas shows in both cases an overall r.m.s. deviation of 0.5 A for the atoms in the main-chain and of 0.4 A for the atoms in the side-chains that make up the hydrophobic core. The iron atom is co-ordinated to four cysteine sulfur atoms forming an almost regular tetrahedron, with Fe-SG distances ranging from 2.27 A to 2.31 A and angles varying from 103 degrees to 115 degrees. The intramolecular hydrogen-bonding pattern is quite comparable to those found in other proteins refined at high resolution. All the polar groups are involved in hydrogen bonds: intramolecular, intermolecular or with solvent molecules. The main structural differences from the other rubredoxins are in the nature and the distribution of some of the charged residues over the molecular surface. The possible influence of several structural factors on the intramolecular and intermolecular electron transfer properties such as the NH...SG bonds, the solvent exposure of the redox center, and the aromatic core is discussed. The conservation, during evolution, of a ring of acidic residues in the proximity of the FeSG4 center suggests that this ring may be implicated in the recognition processes between rubredoxins and their functional partners.

Desulfovibrio↗
PubMed1987

Electrochemical study of the electron exchange between cytochrome c3 and hydrogenase from Desulfovibrio desulfuricans Norway.

The kinetics of the reduction of the Desulfovibrio desulfuricans Norway cytochrome c3 by its physiological partner hydrogenase, in the presence of hydrogen, was investigated by an electrochemical method; from cyclic voltammetry experiments a value of 3 X 10(7) M-1 s-1 was obtained for the second-order rate constant. Results are discussed in terms of specific interactions between physiological partner proteins.

Cytochrome c Group↗
PubMed1987

Structural assignment of the heme potentials of cytochrome c3, using a specifically modified arginine.

In view of the assignment of the four redox potentials values to the four heme groups in the crystallographic structure of Desulfovibrio desulfuricans Norway cytochrome c3, a biochemical approach is reported. A singly modified cytochrome c3 on arginine 73 has been prepared. The study of the redox properties of the modified cytochrome by electrochemistry together with the graphic modelisation of the molecule allow to assign the highest redox potential (-165 mV) to the heme 4 in the three dimensional structure.

Arginine↗
PubMed1987

The evolution of prokaryotic ferredoxins--with a general method correcting for unobserved substitutions in less branched lineages.

Thirty-one bacterial type ferredoxins were examined by means of the parsimony method for their phylogenetic implications. The results show reasonable relationships in that photosynthetic, thermophilic, and desulfovibrio groups are identifiable; but a number of interesting anomalies occur. These include a methanogen sequence that clusters among the desulfovibrios. There are several differences from the phylogeny of Woese. At least two duplications producing paralogous genes are demonstrated, plus the probable existence of two more. The partial internal gene duplication that doubled the length of ferredoxin is confirmed by showing that the probability of the two ancestrally reconstructed halves possessing that much similarity by chance is 10(-7). Howard and co-workers proposed that the two halves of the Azotobacter vinelandii are reversed relative to most other sequences. A phylogeny, drawn with the halves of the azotobacter sequence (and its relatives) reversed produced a tree that had only three less nucleotide substitutions than did the tree without their halves reversed. This plus other evidence suggests that the significantly greater similarity observed across rather than within the halves is more likely the result of convergence.

Amino Acid Sequence↗
PubMed1986

Nucleotide sequence of the promoter and amino-terminal encoding region of the Escherichia coli pepN gene.

The nucleotide sequence of the region probably responsible for regulation of pepN expression and of the region encoding the amino-terminal part of aminopeptidase N, has been determined. The transcription start site was identified by S1 nuclease mapping. All features of the promoter are those of a weak promoter and no obvious structure responsible for regulation was identified, although a possible Pho box is located 63 base pairs upstream from the Pribnow box. The reading frame was unambiguously determined by purifying the protein and by sequencing the first 21 NH2-terminal residues. The NH2-terminal region of aminopeptidase N does not contain any fragment resembling signal sequence and the protein is not produced in a precursor form. A divergent promoter, which might be that of pncB, encoding the nicotinic acid phosphoribosyltransferase (P. Terpstra, personal communication), has also been identified, which allows the assignment of the gene organization in this chromosomal region as ompF asnS pncB pepN.

Amino Acid Sequence↗
PubMed1985

Preliminary 1H-NMR studies of the interaction between cytochrome c3 and ferredoxin I from Desulfovibrio desulfuricans Norway.

The complex formation of two electron transfer proteins, cytochrome c3 and ferredoxin I from Desulfovibrio desulfuricans Norway, has been shown by 1H-NMR spectroscopy. Presence of ferredoxin I produces ferricytochrome c3 1H-NMR spectrum modifications. The chemical shift of perturbated heme methyl resonances has been used to determine the stoichiometry of the complex. At pH 7.6 and 20 degrees C, the two proteins were found to form a complex 1:1 with an association constant, KA, of 10(4) M-1. Two of the four hemes are affected by presence of ferredoxin I and may be involved in the electron transfer sites. The heme methyl resonances are average resonances of free and bound cytochrome c3 resonances, indicating a fast exchange process on the NMR time scale.

Cytochrome c Group↗
PubMed1984

Correlations studies between structural and redox properties of cytochromes C3.

Individual redox potential values are determined for different cytochromes c3. These polyhaemic cytochromes can be divided into two groups: one characterized by only one marked reduction step, the other one giving at least two well-marked reduction steps corresponding to redox potential values ranging from - 0.120 to - 0.400 V. Correlations between potential values and structural data are discussed.

Chemical Phenomena↗
PubMed1984

Electron transfer mechanism and interaction studies between cytochrome C3 and ferredoxin.

Ferredoxin, cytochrome c3 and hydrogenase are specific partners of the sulfate reduction pathway of Desulfovibrio desulfuricans Norway and might be exemplary for electron exchange mechanism studies. Cytochrome c3 contains four low redox potential haems for 13 000 molecular weight. Two ferredoxins isolated from the same bacteria are dimers of 6 000 molecular weight per subunit (Ferredoxin I: one (4 Fe-4S) cluster per subunit, ferredoxin II: two (4 Fe-4 S) clusters per subunit). The amino acid sequence of ferredoxin I is reported and compared to the ferredoxin II sequence. The structural characteristics of ferredoxins and cytochrome c3 should allow a discussion on the nature of the interaction. 1H-NMR spectra of ferredoxin I and cytochrome c3 in the absence and presence of ferredoxin are presented.

Amino Acid Sequence↗
PubMed1983

Coordination of the heme iron in the low-potential cytochromes c-553 from Desulfovibrio vulgaris and Desulfovibrio desulfuricans. Different chirality of the axially bound methionine in the oxidized and reduced states.

The coordination geometry at the heme iron of the cytochromes c-553 from Desulfovibrio vulgaris and Desulfovibrio desulfuricans was investigated by 1H-nuclear magnetic resonance and circular dichroism spectroscopy. Individual assignments were obtained for heme c and the axial ligands. From studies of nuclear Overhauser enhancements the axial histidine imidazole ring orientation relative to the heme group was found to coincide with other c-type cytochromes. In contrast, a new structure was observed for the axial methionine in the reduced cytochromes c-553. This includes S chirality at the iron-bound sulfur atom, but compared to cytochromes c-551 from Pseudomonads and Rhodopseudomonas gelatinosa and cytochrome c5 from Pseudomonas mendocina, which also contain S-chiral methionine, a different spatial arrangement of the gamma- and beta-methylene groups and the alpha carbon of methionine prevails. For the ferricytochromes c-553 R chirality was found for the iron-bound sulfur. This is the first observation of different methionine chirality in different oxidation states of the same c-type cytochrome.

Binding Sites↗
PubMed1983

Comparative studies of monohemic bacterial C-type cytochromes. Redox and optical properties of Desulfovibrio desulfuricans Norway cytochrome C553(550) and Pseudomonas aeruginosa cytochrome C551.

Redox properties of cytochrome c553(550) from Desulfovibrio desulfuricans Norway (Eo' = 0.04 + 0.02 V/NHE) and cytochrome c551 from P. aeruginosa (Eo = 0.25 +/- 0.02 V/NHE) are compared with those of some monohemic c-type cytochromes. The pK value for the equilibrium between the pH-dependent forms of cytochrome c553(550) (pK = 10.3 +/- 0.1) has been also determined. It is to be noted that the difference between redox potentials can extend to nearly 250 mV, though the axial heme ligands are identical. Structural reasons have to be invoked to explain these variations.

Bacterial Proteins↗
PubMed1983

Primary structure of the two (4 Fe-4 S) clusters ferredoxin from Desulfovibrio desulfuricans (strain Norway 4).

The primary structure of a ferredoxin isolated from D. desulfuricans Norway strain, which we called ferredoxin II (Fd II) has been elucidated. This ferredoxin is a dimer constituted of two identical subunits of molecular weight 6000. In ferredoxin II two (4 Fe-4 S) centers are present per subunit instead of one (Fe-S) center as is the case for the other ferredoxins isolated from Desulfovibrio and for Fd I from the same organism. The comparison of amino-acid sequences shows that ferredoxin II presents more homologies with clostridial type ferredoxin than with the ferredoxins from D. gigas and D. africanus.

Amino Acid Sequence↗
PubMed1982

Non-heme iron proteins of Desulfovibrio: the primary structure of ferredoxin I from Desulfovibrio africanus.

Three different ferredoxins have been isolated from the sulfate reducing bacterium, Desulfovibrio africanus. The present paper describes the complete amino acid sequence of D. africanus ferredoxin I. This sequence was determined using automatic protein sequencing in liquid phase and in solid phase. The 61 amino acid residues of the sequence have been aligned with the aid of peptides obtained by cyanogen bromide, cleavage and by tryptic hydrolysis. This ferredoxin which contains 4 cysteine residues represents the most simple case of one (4 Fe-4 S) cluster ferredoxin. A comparison of D. africanus ferredoxin I with D. gigas and Clostridium pasteurianum ferredoxins is presented in terms of structural and possible evolutionary relationships.

Amino Acid Sequence↗