Clusters high-potential iron proteins

G Backes, Y Mino, TM Loehr, TE Meyer, MA Cusanovich, WV Sweeny, ET Adman, J Sand-ers-Loehr. The environment of Ee4S4 clusters in ferredoxms and high-potential iron proteins. New information from X-ray crystallography and resonance Raman spectroscopy. J Am Chem Soc 113 2055-2064, 1991. 

Nonrepetitive but well-defined structures of this type form many important features of enzyme active sites. In some cases, a particular arrangement of coil structure providing a specific type of functional site recurs in several functionally related proteins. The peptide loop that binds iron-sulfur clusters in both ferredoxin and high potential iron protein is one example. Another is the central loop portion of the E—F hand structure that binds a calcium ion in several calcium-binding proteins, including calmodulin, carp parvalbumin, troponin C, and the intestinal calcium-binding protein. This loop, shown in Figure 6.26, connects two short a-helices. The calcium ion nestles into the pocket formed by this structure. 

Three core oxidation states are known for protein-bound [Fe4-S4(S.Cys)4]3+ clusters as illustrated in Figure 2.9. Native proteins exhibit either the [Fe4-S4]2+ + or the [Fe4-S4]3+,2+ redox couple, with proteins involved in the latter couple being referred to historically as HiPIP (high-potential iron protein). The three oxidation states have not been traversed in one protein unless its tertiary structure is significantly perturbed. 

Figure 6.4 Absorption spectrum (A) and CD spectrum (B) of the [Fe4S4] cluster of a high-potential iron protein (HiPIP) from Chromatium sp. (From Cowan, 1997. Reproduced with permission from John Wiley Sons., Inc.)...

Fig. 38. Stereo drawing of the polypeptide backbone of high-potential iron protein. Tight turns are shown with their central peptide as a dark line. The box in the center represents the iron-sulfur cluster.

Figure 10. Resonance Raman spectrum of the 4Fe-AS-4Cys cluster In reduced high-potential Iron protein from Rhodopseudomonas globlformls. Spectral conditions as In Figure 9, but spectrum of frozen solvent not subtracted out. (From Mlno, Y. Loehr,...

The multinuclear tetrahedral iron clusters have the potential for additional formal oxidation states. Because not all of these states have been found in proteins or model compounds, it is possible that some oxidation states may be unstable. For a given Fe S protein only one redox couple is used the other possible states appear to be excluded by restrictions of the protein structure. This selection rule is illustrated with two 4Fe 4S low-molecular-weight electron transfer proteins ferredoxin and high-potential iron protein (HiPIP). The 4Fe 4S clusters in both proteins were shown by X-ray crystallography to be virtually identical. However, the redox potential and oxidation states for the two proteins are vastly... 

High-potential iron proteins, 45 313-314, 344 cluster stability, 45 324-332 function, 45 315-316 residues, 45 322-344 structure and, 45 317-322 redox properties, 45 333-344 solvent accessibility, 45 330, 332-333 source and function, 45 314-316 structure, 45 316-322 hydrogen bonding and, 45 321-322 intermolecular aggregation, 45 322 primary, 45 317-318 secondary and tertiary, 45 318-321... 

Several models have been proposed for the active center of iron and sulphur in Clostridial ferredoxin in which the cysteine residues in the peptide chain participate in the sulphur bridging. Fig 9 166). Unfortunately X-ray analysis of crystals of these proteins has not been completed. It is difficult to confirm that all the irons are clustered in a single linear array 167, 168). X-ray studies of another non-heme iron protein, the high potential iron protein, hipip, from chromatium, carried out by J. Kraut (personal communication), indicate that the four irons of this molecule may be clustered in a tetrahedral array. 

Another group of related electron carriers, the high-potential iron proteins (HIPIP) contain four labile sulfur and four iron atoms per peptide chain 261-266 X-ray studies showed that the 86-residue polypeptide chain of the HIPIP of Chromatium is wrapped around a single iron-sulfur cluster which contains the side chains of four cysteine residues plus the four iron and four sulfur atoms (Fig. 16-15D)261 This kind of cluster is referred to as [4Fe-4S], or as Fe4S4. Each cysteine sulfur is attached to one atom of Fe, with the four iron atoms forming an irregular tetrahedron with an Fe-Fe... 

The functions of the heme is uncertain. The soluble mammalian succinate dehydrogenase resembles closely that of E. coli and contains three Fe-S centers binuclear SI of E° 0 V, and tetranuclear S2 and S3 of -0.25 to -0.40 and + 0.065 V, respectively. Center S3 appears to operate between the -2 and -1 states of Eq. 16-17 just as does the cluster in the Chromatium high potential iron protein. The function of the very low potential S2 is not certain, but the following sequence of electron transport involving SI and S3 and the bound ubiquinone QD-S66 has been proposed (Eq. 18-4). 

Figure 17. Schematic diagrams of some representative topologically chiral proteins.79 (a) Condensed schematic drawing of the L subunit of the quinoprotein TV-MADH. The looped line represents the polypeptide backbone with N and C terminals. Cysteine (or half-cystine) residues are numbered, and their a-carbons are indicated by filled circles. Intrachain disulfide bonds are shown as dashed lines joining a pair of filled circles. The heavy line symbolizes an intrachain cofactor link, (b) Chromatium high potential iron protein (HiPIP), one of several Fe4S4 cluster-containing proteins, (c) Toxin II from the scorpion Androctonus australis Hector. Reprinted with permission from C. Liang and K. Mislow, J. Math. Chem. 1994,15,245. Copyright 1994, Baltzer Science Publishers.

Other biomimetic reactions are based on the catalytic properties of metal ions. Many enzymes require metal ions that function, in one way or another, in oxidation-reduction processes. The wide range of such metal-ion reactions precludes mentioning more than a few in addition to the iron-porphyrin class, and in addition to chlorophyll, a number of enzymes require cobalamin as cofactor ferridoxin and high-potential iron proteins require iron-sulfur clusters, and nitrog-... 

In particular we refer to high-potential iron proteins (HP) and to cubane-like model complexes, which are the only examples of JT effect in biomolecules containing metal clusters we have found in literature " ... 

Mossbauer spectroscopy of Fe-enriched MoFe protein in dithionite-reduced and dye-oxidized oxidation states were interpreted in terms of approximately 50% of the Fe in the protein being present in cubane clusters similar to [4Fe-4S] clusters of simpler Fe-S proteins, e.g., ferredoxins and Chromatium high-potential iron proteins. Spectra of MoFe protein in which P clusters were selectively enriched with Fe were consistent with two of the clusters having slightly different environments. 

High-potential iron proteins (HiPIPs) comprise a subset of the 4Fe-4S cluster family of metalloproteins that are characterized by a positive reduction potential, F , in the range of +50 to +450 mV. This class is differentiated from the 4Fe-4S centers in low-potential ferredoxins, which show a negative E, tjrpically varying between -100 and -650 mV. The origin of these distinct redox properties has been rationalized in terms of the three-state hypothesis of Carter (1), summarized in Scheme 1, and can be attributed to the stability of the common [Fe4S4(SR)4] state. 

Fluorine-labeled analogues of C. vinosum high-potential iron protein have been investigated by F NMR spectroscopy. By incorporation of specific fluorine-labeled amino acid residues, one can insert unique probes at well-defined locations within the protein core. The synthesis and purification of 2-, 3-, and 4-fluorophenylalanine (abbreviated 2-F-, 3-F-, and 4-F-Phe, respectively), 3-fluorotyrosine (3-F-Tyr), and 5-fluorotr3q)tophan (5-F-Trp) derivatives of C. vinosum HiPIP, the assignment of F NMR resonances, the measurement of longitudinal relaxation times, and the temperature dependence of F and resonances have all been reported 42, 43, 136). These measurements were used to examine structural perturbations of mutants, the dynamics of interaction of residues with the cluster, and solvent accessibility, and as a test of the relative contribution of cross-relaxation to magnetization decay. 

The conclusion from these studies is that neither specific aromatic amino acid side chains nor solvent accessibility appear to play a major role in defining the reduction potentials or electron transfer properties of the cluster in high-potential iron proteins. The role of the aromatic core is to maintain a hydrophobic barrier to solvent water and inhibit oxidation and hydrol5 ic degradation of the cluster. 

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