Axial coordination proteins

Each heme unit in myoglobin and hemoglobin contains one ion bound to four nitrogen donor atoms in a square planar arrangement. This leaves the metal with two axial coordination sites to bind other ligands. One of these sites is bound to a histidine side chain that holds the heme in the pocket of the protein. The other axial position is where reversible binding of molecular oxygen takes place. 

The low reactivity of both Cyt111 and Cyt11 toward NO can be attributed to occupation of the heme iron axial coordination sites by an imidazole nitrogen and by a methionine sulfur of the protein (28). Thus, unlike other heme proteins where one axial site is empty or occupied by H20, formation of the nitrosyl complex not only involves ligand displacement but also significant protein conformational changes which inhibit the reaction with NO. However, the protein does not always inhibit reactivity given that Cat and nNOS are more reactive toward NO than is the model complex Fem(TPPS)(H20)2 (Table II). Conversely, the koS values... 

MCD spectroscopy in range 300 to 2000 nm at both ambient and liquid helium (4.2 K) temperatures can yield information about the spin, oxidation, and coordination states of each heme in a multiheme protein such as CCP (75). This technique, in combination with low-temperature X-band EPR spectroscopy, was used to great effect in characterizing the properties of the fully oxidized and MV forms of the P. aeruginosa CCP in solution. At 4.2 K, both hemes in the oxidized enzyme are low-spin ferric, with diagnostic features in the near infrared-MCD (NIR-MCD) spectrum consistent with one heme with His/Met axial coordination and the other with bis-histidine axial coordination this is entirely consistent with the crystal structure. In contrast, at room temperature only the low-potential (bis-histidine coordinated) heme in the C-terminal domain remains completely low-spin, whereas the high-potential (His/Met coordinated) heme exists as mixture of high- and low-spin forms 58). 

Axial Coordination in Nickel Porphyrins and Nickel-Reconstituted Heme Proteins Investigated by Raman-Difference and Transient-Raman Spectroscopy... 

Resonance Raman scattering provides a valuable method of determining the state of axial ligation in nickel-reconstituted heme proteins and Ni-porphyrin complexes. A pattern of shifts in the Raman coresize and oxidation-state marker lines can be used to monitor changes in axial coordination. The shifts in the core-size lines (e.g. indicate an expansion of the core from about 1.96 A for the 4-coordinate Ni porphyrin to 2.04 A for the 6-coordinate species,... 

But, because there was also a first-order term, reduction via a dinitrosyl complex may not be compulsory. It is doubtful that cytochromes could participate in NO reduction via dinitrosyl complexes, because of strong axial coordination of Fe by at least one protein ligand. It is of course possible that the nonheme iron of nitric oxide reductase is the actual site of reduction of NO. 

As in myoglobin, hemoglobin (Fig. 7-23), and cytochrome c (see Fig 16-8), one axial coordination position on the iron of most heme proteins (customarily called the proximal position) is occupied by an imidazole group of a histidine side chain. However, in cytochrome P450 and chloroperoxidase a thiolate (-S ) group from a cysteinyl side chain, and in catalase a phenolate anion from a tyrosyl side chain, occupies the proximal position. The sixth or distal coordination position is occupied by the sulfur atom of methionine in cytochrome c and most other cytochromes with low-spin iron but cytochromes b5 and c3 have histidine. The high-spin heme proteins, such as cytochromes c, ... 

Structural studies95-97,101 103 on cytochromes of the c and c2 types show that the heme group provides a core around which the peptide chain is wound. The 104 residues of mitochondrial cytochrome c are enough to do little more than envelope the heme. In both the oxidized and reduced forms of the protein, methionine 80 (to the left in Fig. 16-8A) and histidine 18 (to the right) fill the axial coordination positions of the iron. The heme is nearly "buried" and inaccessible to the surrounding solvent. 

The development of synthetic enzymes and proteins has also been achieved through the preparation of structurally defined peptide nanostructures. A nice example, reported by DeGrado and co workers [67], is the construction (Fig. 27) of a four a-helix bundle system (72) that was shown to complex four metalloporphyrins by their axial coordination with the imidazole of the properly oriented histidines. This type of structure could be used as an artificial photosynthetic center. Along the same lines, Benson and co-workers [68] recently prepared a miniature hemoprotein, 73, by linking two units of a 13-amino acid peptide to a porphyrin. UV-visible and CD studies confirmed that the metalloporphyrin is indeed sandwiched between the a-helical peptides, as depicted in 73. 

The cobalt center in MeCbl, one of the two important B12 coenzymes, is clearly involved in key steps in catalytic methyl transfer processes. Here, the Co center cycles between Co(I) and Co(III)CH3. In methionine synthase, the proposed mechanism involves direct nucleophilic attack on the C of the Co(III)CH3 group. In model reactions, the thiolate most frequently simply binds tram to the alkyl group to give a product recently established by an x-ray study of a model system. The protein may block access to the Co, thus preventing this reaction common in models. It is likely that the reactive form of the bound cofactor is five-coordinate in the key point in the catalytic cycle. This reactive form will lead to a four-coordinate Co(I) species. The axial coordination of the cofactor by a protein imidazole allows for a finer tuning of the Cbl chemistry and may permit control of the coordination number. Thus, recoordination of Co in the Co(I) state may facilitate attack on methyltetrahydrofolate and re-formation of Co(III)CH3. 

Intriguingly, the blue copper sites, especiaUy those with a carbonyl oxygen at the axial coordination position, display high affinity for Zn + ions. Mutants in which the Met is replaced by Gin or Glu preferentiaUy bind Zn + when expressed in heterologous systems, e.g., Escherichia coli. Examples include azurin, amicyanin, nitrite reductase, and possibly also plastocyanin (Diederix et al., 2000 Hibino et al., 1995 Murphy et al., 1995 Nar et al., 1992a Romero et al., 1993). In the case of azurin it has been shown that both wild-type and the Met—Gin mutant have the same affinity for both Zn +and Cu + (Romero ci a/., 1993). In addition, EXAFS studies showed that some preparations of blue copper proteins purihed from their natural sources also contain small fractions of Zn derivatives (DeBeer George, personal communication). 

The protein s tertiary structure can place any particular atom or group in a suitable position for axial coordination. Thus, the protein folding is responsible for bringing the unusual methionine sulliir atoms (recall that suliiir is a soft donor) in axial sites. This prevents water molecules (which would be the natural choice for Mg, a hard cation) from occupying the axial sites. 

Class II cytochromes c (E° - 100 mV) are found in photosynthetic bacteria, where they serve an unknown function. Unlike their Class I cousins, these c-type cytochromes are high-spin the iron is five-coordinate, with an axial His ligand. These proteins, generally referred to as cytochromes c, are four-a-helix bundles (Figure 6.8). The vacant axial coordination site is buried in the protein interior. 

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