Labile Sulfur Contents

When total iron and non-heme iron in adrenodoxin were determined with o-phenanthroline all the iron existed as non-heme iron and was extractable with 5% trichloroacetic acid. The ratio of iron to protein on a biuret basis was 109 mp. atoms of iron per mg of protein. Based on the dry weight of adrenodoxin, the ratio was 147 mp. atoms of iron per mg protein. The minimum molecular weight per iron atom would be 6,800. 

Assuming a molecular weight of 13,600, there is 2 iron atoms per mole of adrenodoxin. 

In purified adrenodoxin, neither flavin nor heme could be detected. No significant contamination with manganese, cobalt, nickel, copper, zinc, or molybdenum was found in the partially purified preparation by the atomic absorption method as shown in Table 2. 

The iron content of testis non-heme iron protein was found to be 103 mji, atoms per mg of protein on a biuret basis. Although the sedimentation constant is slightly higher than that of adrenodoxin, it would be reasonable to assume that there are 2 iron atoms per mole of protein. 

The analysis for labile sulfur in adrenodoxin is based on the colorimetric determination of methylene blue from p-dimethylamino aniline and Na2S in the presence of FeCls (17). Adrenodoxin was revealed to contain 100 mjj. moles of labile sulfur per mg of protein in a biuret basis. This corresponds to 2 moles of labile sulfur per mole of adrenodoxin. The ratio of iron to labile sulfur is approximately one. 

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The metal and acid labile sulfur contents of these proteins was somewhat lower than expected by comparison with the MoFe proteins. AcP contained 2 0.3 vanadium atoms, 21 1 iron atoms, and 19 0.2 acid-labile sulfur atoms per a2)32 hexamer (172). The a2fi2(S) form of Avl was reported to contain 1.4 0.2 vanadium atoms, 21.4 iron atoms, and 24 acid-labile sulfur ions per molecule, and the afi2 form had lower levels of metal and sulfur (173). 

Samples Iron content Labile sulfur content Activity of steroid 11 (3-hydroxylase... 

A similar result was obtained in reconstitution experiments from heat-treated apoadrenodoxin. A solution (pH 7.4) containing adrenodoxin was allowed to stand for 4.5 hr at 50° C. After the Sephadex G-25 filtration, the colorless sample obtained contained about 90% of the iron present in native adrenodoxin, whereas the labile sulfur content was... 

The bacterial-type iron-sulfur proteins all contain larger amounts of iron and labile sulfide than the plant-type iron-sulfur proteins best estimates for the iron and labile sulfide content being 8 Fe and 8 S per protein molecule (172, 173) for these ferredoxins from Clostridium and from Chromatium. Although these proteins have large amounts of Fe and S, the molecular weights are less than the molecular weights of the... 

The V content of FeVco is variable. The original values reported based on a a 2 structure vary from 0.7 to 2 V atoms, 9 to 23 Fe atoms, and 20 acid-labile sulfur atoms. More recently the V Fe S ratios for different species of A. vinelandii have been given as 1 19 19 for a/32 and 2 30 34 for a-ifa forms. 

Baginsky and Hatefi (155, 156) showed that loss of reconstitution activity appears to be related to a damage in the iron-sulfur system of the enzyme which is not detectable by assay for iron and labile sulfide content. They obtained a preparation of succinate dehydrogenase from complex II which exhibited no reconstitution activity but had an iron labile sulfide flavin ratio close to 8 8 1. They were then able to reactivate this enzyme for reconstitution by treating it with NajS, ferrous ions, and mercaptoethanol, essentially in the same manner as apoferredoxin had been previously converted to ferredoxin (181, 18Z). The reactivated preparation was able to reconstitute with alkali-treated submitochondrial particles or complex II. Analyses showed that the preparation had acquired additional iron and labile sulfide, but control experiments indicated that reconstitution activity was not a spurious effect. The reactiva-... 

The MoFe protein is an aifli tetramer of Mr 220 kDa, and its a and subunits are encoded by the nifD and nifK gene, respectively (Figure 1(a) and Table 1). It contains, in preparations with the highest activity, 2 molybdenum (Mo), 30 to 34 iron (Fe), and an approximately equivalent number of acid-labile sulfur (S ) atoms (Table 1). This metal content is consistent with the presence of two different types of unique metal clusters in the protein, that is, the [8Fe-7S] cluster (P cluster), which is bridged between each a/3 subunit pair, and the [Mo-7Fe-9S-homocitrate] cluster (FeMo cofactor or FeMoco), which is located within each a subunit (Figure la). ... 

The activation mechanism of phosphosulfate linkages (P—O —S)has been studied to understand the chemistry of biological sulfate-transfer reactions of phosphosul-fates of adenosine (APS and PAPS). Several phosphosul-fates were prepared and subjected to several nucleophilic reactions including hydrolysis. In general, phosphosulfates are stable in neutral aqueous mediay but become labile under acidic conditions, resulting in selective S—O fission. This S—O fission appears to occur by unimolecular elimination of sulfur trioxide, which can react with a nucleophilic acceptor, leading to a sulfate-transfer reaction. This process can be accelerated by Mg2+ ion when the solvent is of low water content. Under neutral conditions, divalent metal ions also were found to catalyze nucleophilic reactions, but these occurred on phosphorus to result in exclusive P-O fission. 

DerVartanian et al. (4 ) have examined the EPR properties of the dehydrogenase of Baugh and King (43), which has a flavin iron labile sulfide ratio of 1 28 28. They have concluded that quantitation of the four reduced iron-sulfur centers by double integration accounts for only 36% of the iron content of the preparation. They felt that in this preparation the behavior of the EPR resonances suggests the presence of unidentified iron complexes in addition to iron-sulfur centers. Ohnishi et al. (50) have examined a preparation of complex I made by Ragan and Racker... 

The dehydrogenase preparations obtained by acid-ethanol extraction of particles at elevated temperatures vary considerably in their content of flavin, iron, and labile sulfide, and in their activities. These differences appear to be largely a consequence of destruction of the iron-sulfur chromophore at acid pH. As seen in Fig. 10, incubation of the low molecular weight dehydrogenase preparation of Hatefl and Stempel at pH 4.8 and 38° resulted after 1 hr in nearly complete loss of labile sulfide (Fig. IOC) and reductase activity with respect to menadione, cytochrome c. 

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