Rubredoxin from Clostridium pasteurianum

The crystallographic structure of rubredoxin from Clostridium pasteurianum at 2.5 A, a resolution sufficient to reveal the sequence of several of the bulky amino acid side chains, shows the iron coordinated to two pairs of cysteine residues located rather near the termini of the polypeptide chain (Fig. 1). A related rubredoxin, with a three times larger molecular weight, from Pseudomonas oleovorans is believed to bind iron in a similar fashion. This conclusion is based on physical probes, especially electron paramagnetic resonance spectroscopy, all of which indicate that the iron is in each case situated in a highly similar environment however, the proteins display some specificity in catalytic function. 

The simplest of these proteins are rubredoxins, which are bacterial proteins having a characteristic red colour (from which their name is derived) containing an FeS4 assembly, consisting of an Fe(III) ion coordinated to four cysteine groups. The typical tetrahedral structure of this group is illustrated in Figure 17 for the rubredoxin isolated from Clostridium pasteurianum (FW 6100).35... 

Figure 17 X-Ray structure of the active site of the rubredoxin from Clostridium pasteurianum...

Figure 18 Cyclic voltammograms recorded at an edge-oriented pyrolitic graphite electrode in an aqueous solution (pH 8) of the rubredoxin from Clostridium pasteurianum. In the absence (a) and in the presence (b) of [Cr(NH3)6J3+. Scan rate 0.02 V s J...

Rubredoxins. Rubredoxins are the simplest form of iron-sulfur proteins in which iron is bound to the sulfur atom of cysteine as shown in Fig. lA. One of the first rubredoxins isolated was from an anaerobic bacterium, Clostridium pasteurianum, by Lovenberg and Sobel (9). The protein is composed of 54 amino acids and has a molecular weight of 6,000. The oxidized form has absorbance maxima at 380 and 490 nm. The biological role of the rubredoxin isolated from C. pasteurianum is stiU unknown. 

Figure 16-16 (A) Superimposed stereoscopic a-carbon traces of the peptide chain of rubredoxin from Clostridium pasteurianum with either Fe3+ (solid circles) or Zn2+ (open circles) bound by four cysteine side chains. From Dauter et al.27i (B) Alpha-carbon trace for ferredoxin from Clostridium, acidurici. The two Fe4S4 clusters attached to eight cysteine side chains are also shown. The open circles are water molecules. Based on a high-resolution X-ray structure by Duee et al.267 Courtesy of E. D. Duee. (C) Polypeptide chain of a chloroplast-type ferredoxin from the cyanobacterium Spirulina platensis. The Fe2S2 cluster is visible at the top of the molecule. From Fukuyama et al.276 Courtesy of K. Fukuyama.

Fig. 5.15. The iron core of rubredoxin from Clostridium pasteurianum.

Dauter Z, Wilson KS, Sieker LC, Moulis JM, Meyer J (1996) Zinc- and iron-rubredoxins from Clostridium pasteurianum at atomic resolution A high-precision model of a ZnS4 coordination unit in a protein. Proc Natl Acad Sci USA 93 8836-8840... 

Xiao ZG, Maher MJ, Cross M, Bond CS, Guss JM, Wedd AG (2000) Mutation of the surface valine residues 8 and 44 in the rubredoxin from Clostridium pasteurianum solvent access versus structural changes as determinants of reversible potential. J Biol Inorg Chem 5 75-84... 

Rubredoxin was first described in 1963 by Buchanan [1], and in 1964 by Mortensen [2] during the purification of ferredoxin from Clostridium pasteuriaman. In 1965, Lovenberg and Sobel [3] were able to isolate rubredoxin, a small protein that could replace ferredoxin as an electron carrier in reactions mediated by extracts from C. pasteurianum, and named it rubredoxin (from ruber — red). The chemical feature which, historically, has distinguished rubredoxins from other iron-sulfur proteins. 

Lovenberg, W., Williams, W. M., Further Observations on the Chemical Nature of Rubredoxin from Clostridium pasteurianum/ Biochemistry (1969) 8,141. 

Rubredoxins (Rd), small iron-containing proteins occurring in various sulfur-metabolizing bacteria and archea involved in electron transfer. Sometimes Rd are classified as iron-sulfur proteins. However, they do not contain inorganic sulfide. Rubredoxin from Clostridium pasteurianum (Mr 6 kDa 52-54 aa) shows functional similarity to ferredoxin but contains, in contrast to the latter, only one iron bound by the sulfur atoms of four cysteine residues. The iron-sulfur complex shows tetrahedral symmetry. The 3-D structures of a number of rubredoxins have been solved. The folds belong to the a+ fi class, with two a-helices and two to three /3-strands [W. Lovenberg, B. E. Sobel, Proc. Natl. Acad. Sci. USA 1965, 54,193 M. Frey etal.,J. Mol. Biol. 1987, 197, 525]. 

The rubredoxins (Rd) contain the simplest of the Fe-S structures, with a single iron ion surrounded tetrahedrally by four Fe-S(Cys) bonds. " - The RR spectrum of Rd from Clostridium pasteurianum was one of the first RR spectra obtained for a biological molecule. " As illustrated in Fig. 19, this spectrum could be interpreted as arising from a simple tetrahedral complex indeed, it resembled closely the spectrum of the isoelec-tronic cluster [FeCl4]". Further studies, with better resolution and a wider range of excitation wavelengths, have revealed considerable com-... 

One of the simplest of the non-haem iron proteins is the rubredoxin from Clostridium pasteurianum, which contains one iron atom per molecule and undergoes a reversible one-electron oxidation and reduction. The... 

Fig. 29.14 (a) A ribbon representation of the metalloprotein rubredoxin from the bacterium Clostridium pasteurianum. The position of the Fe atom in the active site is shown, (b) Detail of the active site showing the tetrahedral arrangement of the Cys residues that bind the Fe centre. Flydrogen atoms are omitted colour code Fe, green S, yellow C, grey. 

Rubredoxin a redoxin, functionally similar to Fer-redoxin (see). M, 6,000. R. was isolat from Clostridium pasteurianum synthesis of R. appears to be promoted by a relative deficien<7 of iron. It contains one Fe atom/molecule of protein, which is less than the Fe content of ferredoxin. Under acid conditions it is more stable than ferredoxin, and it has a more positive redox potential (E o -0.057 V) thus when R. replaces ferredoxin in a feiredoxin-dependent reaction the reaction rate is decreased. The iron is bound by coordination with 4 cysteinyl residues other possible ligands are tyrosine and lysine. Redoxins similar to R. have been isolated from Peptostreptococcus elsde-nii and other baeteria. R. from Micrococcus aerogenes contains 53 amino acid residues of known sequence. 

An intense eifort by physicists, physical chemists, biochemists, inorganic chemists and X-ray crystallographers has now defined the common attribute of the nonhaem iron proteins. It is an iron centre tetrahedrally coordinated by four sulphur ligands. A number of variations within this theme are recognized. The simplest case is found to be rubredoxin from Clostridium pasteurianum, a 6000 molecular weight protein whose exact electron transport function is unknown. 

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