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C Veeger

Publications and source records attributed to C Veeger.

At least 73 records · Page 4Linked to original sources
PubMed1987

Time-resolved fluorescence and circular dichroism of porphyrin cytochrome c and Zn-porphyrin cytochrome c incorporated in reversed micelles.

Interactions between fluorescent horse heart cytochrome c derivatives (e. g. porphyrin cytochrome c and Zn-porphyrin cytochrome c) with surfactant interfaces in reversed micellar solutions have been studied, using different spectroscopic techniques. Anionic [sodium bis(2-ethylhexyl)sulfosuccinate, AOT] and cationic (cetyltrime-thylammonium bromide, CTAB) surfactant solutions have been used in order to investigate the effects of charge interactions between proteins and interfaces. Circular dichroism reveals that much of the protein secondary structure is lost in AOT-reversed micelles, especially when the molar water/surfactant ratio, wo, is high (wo = 40), whereas in CTAB-reversed micelles secondary structure seems to be preserved. Time-resolved fluorescence measurements of the porphyrin in the cytochrome c molecule yields information about the changes in structure and the dynamics of the protein upon interaction with surfactant assemblies both in aqueous and in hydrocarbon solutions. With AOT as surfactant a strong interaction between protein and interface can be observed. The effects found in aqueous AOT solution are of the same kind as in hydrocarbon solution. In the CTAB systems the interactions between protein and surfactant are much less pronounced. The measured effects on the fluorescence properties of the proteins are different in aqueous and hydrocarbon solutions. In general, the observations can be explained by an electrostatic attraction between the overall positively charged protein molecules and the anionic AOT interface. Electrostatic attraction can also occur between the cytochrome c derivatives and CTAB because there is a negatively charged zone on the surface of the proteins. From the fluorescence anisotropy decays it can be concluded that in the CTAB-reversed micellar system these interactions are not important, whereas in an aqueous CTAB solution the proteins interact with surfactant molecules.

Circular Dichroism↗
PubMed1987

Spectroscopic properties of horse liver alcohol dehydrogenase in reversed micellar solutions.

Catalytic and spectroscopic properties of alcohol dehydrogenase from horse liver, incorporated in reversed micellar media, have been studied. Two different reversed micellar systems have been used, one containing an anionic [sodium bis(2-ethylhexyl)sulfosuccinate, AOT], the other containing a cationic (cetyltrimethylammonium bromide, CTAB) surfactant. With 1-hexanol as substrate the turnover number of the enzyme in AOT-reversed micelles is strongly dependent on the water content of the system. At low wo ([H2O]/[surfactant]) (wo less than 20) no enzymatic activity can be detected whereas at high wo (wo = 40) the turnover is only slightly lower than in aqueous solution. In CTAB-reversed micelles the dependence of the turnover number on wo is much less. The enzymatic activity is in this case significantly lower than in aqueous solution and increases only slightly with an increasing water content of the reversed micelles. Possible interactions of the protein with the surfactant interfaces in the reversed micellar media were studied via circular dichroism and fluorescence measurements. From the circular dichroism of the protein backbone it is observed that the protein secondary structure is not significantly affected upon incorporation in the reversed micelles since the far-ultraviolet spectrum is not altered. Results from time-resolved fluorescence anisotropy experiments indicate that, especially in AOT-reversed micelles, interactions between the protein and the surfactant interface are largely electrostatic in nature, as evident from the dependence on the pH of the buffer used. In CTAB-reversed micellar solutions such interactions appear to be much less pronounced than in AOT.

Alcohol Dehydrogenase↗
PubMed1987

Binding of ADP and orthophosphate during the ATPase reaction of nitrogenase.

The pre-steady-state ATPase activity of nitrogenase from Azotobacter vinelandii was investigated. By using a rapid-quench technique, it has been demonstrated that with the oxidized nitrogenase complex the same burst reaction of MgATP hydrolysis occurs as observed with the reduced complex, namely 6-8 mol orthophosphate released/mol MoFe protein. It is concluded that the pre-steady-state ATPase activity is independent of electron transfer from Fe protein to MoFe protein. Results obtained from gel centrifugation experiments showed that during the steady state of reductant-independent ATP hydrolysis there is a slow dissociation of one molecule of MgADP from the nitrogenase proteins (koff less than or equal to 0.2 s-1); the second MgADP molecule dissociates much faster (koff greater than or equal to 0.6 s-1). Under the same conditions orthophosphate was found to be associated with the nitrogenase proteins. The rate of dissociation of orthophosphate from the nitrogenase complex, as estimated from the gel centrifugation experiments, is in the same order of magnitude as the steady-state turnover rate of the reductant-independent ATPase activity (0.6 mol Pi formed X s-1 X mol Av2(-1) at 22 degrees C). These data are consistent with dissociation of orthophosphate or MgADP being rate-limiting during nitrogenase-catalyzed reductant-independent ATP hydrolysis.

Adenosine Diphosphate↗
PubMed1987

Purification and characterization of Desulfovibrio vulgaris (Hildenborough) hydrogenase expressed in Escherichia coli.

Hydrogenase from Desulfovibrio vulgaris (Hildenborough) is a heterologous dimer of molecular mass 46 + 13.5 kDa. Its two structural genes have been cloned on a 4664-base-pair fragment of known sequence in the vector pUC9. Expression of hydrogenase polypeptides in Escherichia coli transformed with this plasmid is poor (approximately 0.1% w/w of total protein). Deletion of up to 1.9 kb of insert DNA brings the gene encoding for the large subunit in close proximity to the lac promotor of pUC9 and results in a 50-fold increased expression of hydrogenase polypeptides in E. coli. The protein formed is inactive and was purified in order to delineate its defect. Complete purification was achieved with a procedure similar to that used for the isolation of active hydrogenase from D. vulgaris H. The derived protein is also an alpha beta dimer and electron-paramagnetic resonance studies indicate the presence of the electron-transferring ferredoxin-type iron-sulfur clusters. In contrast to the native protein from D. vulgaris H, these can only be reduced with dithionite, not with hydrogen, indicating that the hydrogen-binding active centre which also contains an iron-sulfur cluster is missing.

DNA Transposable Elements↗
PubMed1986

EPR of a novel high-spin component in activated hydrogenase from Desulfovibrio vulgaris (Hildenborough).

The EPR of reoxidized hydrogenase from Desulfovibrio vulgaris (H.) has been reinvestigated. In contrast to other workers [(1984) Proc. Natl. Acad. Sci. USA 81, 3728-3732] we find the axial signal with g = 2.06; 2.01 to be only a minor component of concentration 0.03 spin/mol. In the spectrum of fully active reoxidized enzyme this signal is overshadowed by a rhombic signal (0.1 spin/mol) with g = 2.11; 2.05; 2.00 reminiscent of the only signal found for other oxidized bidirectional hydrogenases. In addition, a novel signal has been detected with geff = 5.0 which, under the assumptions that S = 2 and [delta ms] = 2, quantitates to roughly one spin/mol. Ethylene glycol affects the relative intensity of the different signals. It is suggested that O2 sensitization parallels a spin-state transition of an iron-sulfur cluster.

Desulfovibrio↗
PubMed1986

Characterization of three different flavodoxins from Azotobacter vinelandii.

The flavodoxins from Azotobacter vinelandii cells grown N2-fixing and from cells grown on NH4OAc have been purified and characterized. The purified flavodoxins from these cells are a mixture of three different flavodoxins (Fld I, II, III) with different primary structures. The three proteins were separated by fast protein liquid chromatography; Fld I eluted at 0.38 M KCl, Fld II at 0.43 M KCl and Fld III at 0.45 M KCl. The most striking difference between the three flavodoxins was the midpoint potential (pH 7.0, 25 degrees C) of the semiquinone/hydroquinone couple, which was -320 mV for Fld I and -500 mV for the other two flavodoxins (Fld II and Fld III). All three flavodoxins were present in cells grown on NH4OAc. In cells grown on N2 as N source only Fld I and Fld II were found. The concentration of Fld II was 10-fold higher in N2-fixing cells than in cells grown on NH4OAc. Evidence has been obtained that Fld II is involved in electron transport to nitrogenase. As will be discussed, our observation that preparations of Azotobacter flavodoxin are heterogeneous, has consequences for the published data.

Azotobacter↗
PubMed1986

Studies on the mechanism of electron transport to nitrogenase in Azotobacter vinelandii.

The involvement of the cytoplasmic membrane in electron transport to nitrogenase has been studied. Evidence shows that nitrogenase activity in Azotobacter vinelandii is coupled to the flux of electrons through the respiratory chain. To obtain information about proteins involved, the changes occurring in A. vinelandii cells transferred to nitrogen-free medium after growth on NH4Cl (depression of nitrogenase activity) were studied. Synthesis of the nitrogenase polypeptides was detectable 5 min after transfer to nitrogen-free medium. No nitrogenase activity could be detected until t = 20 min, whereupon a linear increase of nitrogenase activity with time was observed. Synthesis of nitrogenase was accompanied by synthesis of flavodoxin II and two membrane-bound polypeptides of Mr 29,000 and 30,000. Analysis with respect to changes in membrane-bound NAD(P)H dehydrogenase activities revealed the induction of an NADPH dehydrogenase activity, which was not detectable in membranes isolated from cells grown in the presence of NH4OAc. This induced activity was associated with the appearance of a polypeptide of Mr 29,000 in the NADPH dehydrogenase complex.

Azotobacter↗
PubMed1986

Application of hydrogenase in biotechnological conversions.

Evidence will be presented in this review article that the application of hydrogenase has large biotechnological possibilities. Our investigations show: Fast reaction of hydrogenase at an electrode surface to reduce H+; Photochemical production of H2 by hydrogenase by photosensitized Ru-complexes dissolved in reversed micellar membranes and vectorial H+ transport through the membrane to the water phase; The production of fine chemicals in reversed micelles by a system containing specific enzymes, hydrogenase and H2. The rules to obtain maximal conversion rates with this system will be presented.

Catalysis↗
PubMed1985

Properties of the MgATP and MgADP binding sites on the Fe protein of nitrogenase from Azotobacter vinelandii.

Flow dialysis was used to study the binding of MgATP and MgADP to the nitrogenase proteins of Azotobacter vinelandii. Both reduced and oxidized Av2 bind two molecules of MgADP, with the following dissociation constants: reduced Av2, K1 = 0.091 +/- 0.021 mM and K2 = 0.044 +/- 0.009 mM; oxidized Av2, K1 = 0.024 +/- 0.015 mM and K2 = 0.039 +/- 0.022 mM. Binding of MgADP to reduced Av2 shows positive co-operativity. Oxidized Av2 binds two molecules of MgATP with dissociation constants K1 = 0.049 +/- 0.016 mM and K2 = 0.18 +/- 0.05 mM. Binding data of MgATP to reduced Av2 can be fitted by assuming one binding site, but a better fit was obtained by assuming two binding sites on the protein with negative co-operativity and with dissociation constants K1 = 0.22 +/- 0.03 mM and K2 = 1.71 +/- 0.50 mM. It was found that results concerning the number of binding sites and the dissociation constants of MgATP-Av2 and MgADP-Av2 complexes depend to a great extent on the specific activity of the Av2 preparation used, and that it is difficult to correct binding data for inactive protein. No binding of MgADP to Av1 could be demonstrated. Binding studies of MgADP to a mixture of Av1 and Av2 showed that Av1 did not affect the binding of MgADP to either oxidized or reduced Av2. Inhibition studies were performed to investigate the interaction of MgATP and MgADP binding to oxidized and reduced Av2. All the experimental data can be explained by the minimum hypothesis, i.e. the presence of two adenine nucleotide binding sites on Av2. MgATP and MgADP compete for these two binding sites on the Fe protein.

Adenosine Diphosphate↗
PubMed1985

A study of one of the iron-sulphur clusters in oxidized hydrogenase from Megasphaera elsdenii by magnetic-circular-dichroism spectroscopy.

The m.c.d. spectrum of the oxidized state of hydrogenase from Megasphaera elsdenii has been measured at liquid-helium temperatures. This oxidation state of the enzyme displays a characteristic rhombic e.p.r. signal with g-values of 2.101, 2.052 and 2.005 assigned previously to a [4Fe-4S]3+ cluster as in oxidized HiPIP (high-potential iron-sulphur protein) [Van Dijk, Grande, Mayhew & Veeger (1980) Eur. J. Biochem. 107, 251-261]. The low-temperature m.c.d. spectrum shows no features attributable to an oxidized four-iron cluster of the HiPIP type, but does reveal broad, positive peaks at 460 and 730 nm, which magnetize in a manner untypical of a spin S = 1/2 cluster with g-values close to 2. The m.c.d. spectrum is most closely similar to that of dye-oxidized P-clusters known in the enzyme nitrogenase. It is therefore proposed that the rhombic e.p.r. spectrum at a g-value close to 2 arises from an m.c.d.-silent radical species that may be related chemically to the cysteine persulphide species, RS-S., recently found in the hexacyanoferrate-oxidized seven-iron ferredoxin of Azotobacter vinelandii [Morgan, Stephens, Devlin, Stout, Melis & Burgess (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 1931-1935].

Circular Dichroism↗
PubMed1985

The catalytic activity of nitrogenase in intact Azotobacter vinelandii cells.

The influence of the growth conditions on the concentration of nitrogenase and on the nitrogenase activity, was studied in intact Azotobacter vinelandii cells. It was observed that whole cell nitrogenase activity could be enhanced in two ways. An increase of the growth rate of cells was accompanied by an increase in whole cell nitrogenase activity and by an increase in the concentration of nitrogenase in the cells. The molar ratio of Fe protein:MoFe protein was 1.47 +/- 0.17 and independent of the growth rate. Activity measurements in cell extracts showed that the catalytic activity of the nitrogenase proteins was independent of the growth rate of cells. The second way to increase whole cell nitrogenase activity was to expose cells to excess oxygen. Whole cells were exposed for 2.5 h to an enhanced oxygen-input rate. After this incubation nitrogenase activity was increased without an increase in protein concentration. It is calculated that the catalytic activity of the Fe protein in these cells was 6200 nmol C2H4 formed X min-1 X (mg Fe protein)-1. With these cells and with cells grown at a high growth rate, 50% of the whole cell activity is lost by preparing a cell-free extract. It will be demonstrated that this inactivation is partly caused by the activity measurements in vitro. When dithionite was replaced by flavodoxin as electron donor, a maximal catalytic activity of 4500 nmol C2H4 formed X min-1 X (mg Fe protein)-1 was measured in vitro for the Fe protein. The results are discussed in relation to the present model for nitrogenase catalysis.

Azotobacter↗
PubMed1984

Rules for the regulation of enzyme activity in reserved micelles as illustrated by the conversion of apolar steroids by 20 beta-hydroxysteroid dehydrogenase.

20 beta-Hydroxysteroid dehydrogenase was enclosed in reversed micellar media consisting of cetyltrimethyl-ammonium bromide, hexanol, organic solvent and Hepes buffer. The influence of the composition of these media on the enzymatic reduction of the apolar steroids progesterone and prednisone was investigated by varying the water content, concentration of hexanol and type of organic solvent. By changing the water content and the type of organic solvent, the hexanol to cetyltrimethylammonium bromide ratio in the interphase can be varied. This ratio was determined by phase boundary titrations. It was found that the higher this ratio, the higher the rate of steroid conversion. From variations of the hexanol content it was concluded that the rate of steroid conversion is determined by the hydrophobicity of the steroid relative to the hydrophobicity of the continuous phase and the hydrophobicity of the interphase. The hydrophobicity of the phases was expressed in log P-values. Log P is defined as the logarithm of the partition coefficient in an octanol-water two-phase system. This enabled us to derive the following relations between the hydrophobicity values for the substrate (log Ps), for the interphase (log Pi) and for the continuous phase (log Peph): [log Pi-log Ps] must be minimal to ensure a high steroid concentration in the interphase and [log Pcph-log Ps] must be large to keep the steroid concentration in the continuous phase low. With these considerations, for any given apolar compound, a medium can be composed that gives optimal enzymatic conversion.

20-Hydroxysteroid Dehydrogenases↗
PubMed1984

The composition of the pyruvate dehydrogenase complex from Azotobacter vinelandii. Does a unifying model exist for the complexes from gram-negative bacteria?

An improved purification procedure of the pyruvate dehydrogenase complex of Azotobacter vinelandii is described. This procedure minimizes losses of components and results in the isolation of the pure complex with a specific activity of 15-19 U/mg and an overall yield of 40%. The chain ratio of the three components was determined by covalent modification of the lysine residues with trinitrobenzene sulfonic acid, followed by separation of the components on sodium dodecyl sulfate gels. These determinations yielded an average chain ratio of 1.3:1:0.5 for E1:E2:E3 respectively. Based on E2 this corresponds with a minimum molecular mass of approximately 216 kDa. Because the molecular mass of the complex has been determined previously to be 800 +/- 50 kDa, it is concluded that the complex as isolated from A. vinelandii is based on a tetramer of E2 chains. The complex can be resolved into its individual components, which can be recombined to yield a fully active complex. Titration of E2E3 subcomplexes with E1 resulted in maximum complex activity at an E1/E2 ratio of 1.5-1.6. Similar titrations of E1E2 subcomplexes with E3 resulted in maximum activity at an E3/E2 ratio of 0.45-0.55. From these experiments it is concluded that the complex has maximum activity with a composition of three E1 dimers, one E2 tetramer and one E3 dimer. With excess of either E1 or E3 a decrease in activity is observed which indicates competition between these components for binding sites on E2. As shown before [Bosma, H.J., de Kok, A., Markwijk, B.W., and Veeger, C. (1984) Eur. J. Biochem. 140, 273-280], the isolated E2 component is composed of 32 peptide chains of 66 kDa each. Upon addition of E1 or E3, E2 dissociated into tetramers. Dissociation is complete upon the addition of four E1 dimers of four E3 dimers per E2 tetramer. Addition of E1 to saturated E2E3 subcomplex or E3 to saturated E1E2 subcomplex did not result in extra binding but rather in displacement of bound E3 or E1 respectively. It is therefore concluded that the binding sites of E1 and E3 to the E2 chains are either identical or so closely spaced that steric hindrance prevents simultaneous binding of both components. A model is presented based on the cubic structure of the isolated E2 component. In this model the 32 E2 peptide chains are arranged in tetramers in the corners of the cube. This model is discussed in connection with the existing model for the Escherichia coli complex.

Amino Acids↗
PubMed1984

The size of the pyruvate dehydrogenase complex of Azotobacter vinelandii. Association phenomena.

Sedimentation analysis and light-scattering studies indicate that the aggregation state of the pyruvate dehydrogenase complex of Azotobacter vinelandii in 50 mM potassium phosphate (pH 7.0) can be described in terms of a monomer-dimer equilibrium with a dissociation constant of 6.8 microM. The apparent molecular mass of the monomeric particle is 750 000-850 000 Da. The equilibrium is shifted to the monomeric species when pressure is applied on the system. Pressure-jump experiments yielded a relaxation time of about 70 ms. In the presence of 3% poly(ethylene glycol) 6000 and 10 mM MgCl2, further association takes place to a system that can be described in terms of dimer-tetramer-octamer equilibria. Upon applying a pressure of 80 MPa to this system these equilibria are shifted to the dimeric state but some monomer formation cannot be excluded. Release of pressure shows that the relaxation time of the dimer-tetramer equilibrium is less than 5 s, that of tetramer-octamer equilibrium is of the order of minutes. The isolated E2 component has a molecular mass of 2 000 000 +/- 100 000 Da; and thus consists of about 30 E2 peptide chains. Electron micrographs are similar to those of the E2 component of the Escherichia coli complex, which were interpreted as cubic structures with an octagonal symmetry. Upon addition of E1 to the pure E2 component, changes in the assembly occur and mixtures of large (E. coli-like, 22-45 S) and small (A. vinelandii-like, 11-18 S) subcomplexes are obtained. The two forms of the subcomplexes are in slow equilibrium (relaxation time 10-30 min). It is proposed that the E2 tetramer of the intact pyruvate dehydrogenase complex of A. vinelandii is represented by the corner structures of the isolated E2 component.

Azotobacter↗
PubMed1983

On the formation of an oxygen-tolerant three-component nitrogenase complex from Azotobacter vinelandii.

Conditions are defined in which the oxygen-labile nitrogenase components from Azotobacter vinelandii can be protected against oxygen inactivation by the so-called Fe/S protein II. It is demonstrated that oxygen protection can be achieved by complex formation of the three proteins. Complex formation was studied by gel chromatography. Only when the three proteins are in the oxidized state and MgCl2 is present, can an oxygen-tolerant complex be isolated. Quantitative SDS/polyacrylamide gel electrophoresis of such complexes, yielded an average ratio of nitrogenase component 2/nitrogenase component 1 (Av2/Av1) of 2.4 +/- 0.5. Protection by Fe/S protein II was correlated with the amount of [2 Fe-2S] clusters present in the protein and not by the amount of protein. Measurements of the amount of iron and sulfide of Fe/S protein II showed that the iron and sulfide content of the protein was variable. The maximum values found indicate that Fe/S protein II contains two [2Fe-2S] clusters per dimer of 26 kDa. Full protection by Fe/S protein II was obtained with a ratio of Fe/S protein II/Av1 of 1.1 +/- 0.2; the Fe/S protein II containing two [2Fe-2S] clusters per dimer of 26 kDa. When Fe/S protein II contains less [2Fe-2S] clusters, more protein is necessary to obtain full protection. The three-component nitrogenase complex is also oxygen stable in the presence of MgATP or MgADP. Analysis in the ultracentrifuge showed that the major fraction of the reconstituted complex has a sedimentation coefficient centered around 34S. A small fraction (less than 30%) sediments with values centered around 111 S. This suggests an average mass for the oxygen-stable nitrogenase complex of 1.5 MDa. Taking into account the determined stoichiometry of the individual proteins, the molecular composition of the oxygen-stable nitrogenase complex is presumably 4 molecules of AV1,8--12 molecules of aAV2 and 4--6 molecules of Fe/S protein II containing two [2Fe-2S] clusters per dimer of 26 kDa.

Azotobacter↗
PubMed1983

A pulse-radiolysis study of the reduction of flavodoxin from Megasphaera elsdenii by viologen radicals. A conformational change as a possible regulating mechanism.

Pulse-radiolysis experiments were performed on solutions containing methyl or benzyl viologen and flavodoxin. Viologen radicals are formed after the pulse. The kinetics of the reaction of these radicals with flavodoxin were studied. The kinetics observed depend strongly on the concentration of oxidized viologen. Therefore one must conclude that a relatively stable intermediate is formed after the reduction of flavodoxin. The midpoint potential of the intermediate state is -(480 +/- 30) mV, and is hardly dependent on the pH between 7 and 9.2. Due to a conformational change (k2 approximately equal to 10(5)S-1) the intermediate state decays to the stable semiquinone form of flavodoxin. The delta G of the conformational change at pH 8 is about 29 kJ mol -1 (0.3 eV). This means that the upper limit for the pK of N-5 in the semiquinone form will be 13. The activation energy of the conformational change is 43 kJ mol -1 (0.45 eV). The reaction between methyl viologen radicals and the semiquinone of flavodoxin can be described by a normal bimolecular reaction. The reaction is diffusion-controlled with a forward rate constant of (7 +/- 1) X 10(8) M -1S -1 (pH 8, I = 55 mM). The midpoint potential of the semiquinone/hydroquinone was found to be -(408 +/- 5) mV. A consequence of the intermediate state is that flavodoxin (Fld) could be reduced by a two-electron process, the midpoint potential of which should be located between -440 mV less than Em (Fld/FldH-) less than -290 mV. The exact value will depend on the delta G of the conformational change between the fully reduced flavodoxin with its structure in the oxidized form and the fully reduced flavodoxin with its structure in the hydroquinone form. The conditions are discussed under which flavodoxin could behave as a two-electron donor.

Electron Transport↗
PubMed1983

Fully active Fe-protein of the nitrogenase from Azotobacter vinelandii contains at least eight iron atoms and eight sulphide atoms per molecule.

The Fe-protein of the Azotobacter vinelandii nitrogenase enzyme complex contains a variable iron and sulphide content. The iron and sulphide content of the protein is dependent upon the specific activity. Up to a specific activity of 1000 nmol C2H4 produced X min-1 X mg Av-1(2), three iron and three sulphide atoms per molecule Av2 are found. At specific activities above 1000 nmol C2H4 produced X min-1 X mg Av-1(2), a linear relationship between specific activity and iron and sulphide content of Av2 is found. The maximum values found are 8.8 iron atoms and 8.6 sulphide atoms/molecule at a specific activity of 2250 nmol C2H4 produced X min-1 X mg Av-1(2). Also the experimental molar absorption coefficients at 430 nm of the oxidized and reduced forms depend on the specific activity. The highest values found are 15.9 mM-1 cm-1 and 9.1 mM-1 cm-1, respectively. Since occasionally the preparations with specific activities around 3000 nmol X min-1 X mg-1 are isolated which contain more than 10 iron atoms and 11 sulphide atoms per molecule, it cannot be excluded that under certain physiological conditions Av2 contains even more than two [4 Fe-4 S] clusters. The addition of MgATP induces a conformational change in the Fe-protein which results in a higher reactivity with iron chelators. But irrespective of the specific activity, the amount of iron extracted from the protein after addition of MgATP never exceeds four atoms/molecule. The results are discussed with respect to the present molecular model of the Fe-protein.

Azotobacter↗