Trans effects on cysteine ligation in the proximal HIS93CYS variant of horse heart myoglobin

Hildebrand, Dean P.; Tang, Hai-Lun; Smith, Michael L.; Mauk, A. Grant

doi:10.1016/0162-0134(95)97525-u

Cited by 5 publications

(7 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…One possibility is that, when the heme in wild-type LmPP is reduced, the cysteine (thiolate) distal bond may be broken and the subsequently reduced state of wild-type LmPP can display a high spin with histidine ligated. As seen for wild-type LmPP, earlier ligand switching is reported in many heme proteins that are cysteinate-bound in the ferric state and do not retain such coordination in the ferrous state, like the inactive P420 forms of P450s, 43 and the inactive C420 form of CPO, 44 the CO-sensing CooA protein, 10,55 proximal His-to-Cys mutants such as that of myoglobin, 56 the CCP mutant, 34 and heme oxygenase. 57 We have conducted mutational studies in LmPP and have made a possible schematic diagram of axial ligands of ferric and ferrous forms (Figure 7).…”

Section: ■ Discussionmentioning

confidence: 70%

Identification of Proximal and Distal Axial Ligands in Leishmania major Pseudoperoxidase

et al. 2013

View full text Add to dashboard Cite

Previous optical and electron paramagnetic resonance (EPR) spectroscopic studies of the newly discovered peroxynitrite scavenging pseudoperoxidase from Leishmania major (LmPP) suggested that ferric LmPP contained a six-coordinate low-spin (6cLS) heme with a thiolate ligand, presumably a cysteine, bound to its heme iron. To identify the axial ligands of LmPP, we exploit a systematic mutational analysis of potential heme ligands. On the basis of UV-visible and EPR spectroscopy, we report that the substitution of the proximal His206 with alanine in LmPP alters the 6cLS to a five-coordinate high spin (5cHS) form at pH 4.0 that has a spectrum characteristic of a Cys-ligated 5cHS derivative. The electronic absorption and EPR analysis of all alanine-substituted Cys and Met single mutants establish that when Cys107 is replaced with alanine, a new species appears that has a spectrum characteristic of a histidine-ligated 5cHS derivative at pH 4.0. Together, these results suggest that His206 and Cys107 act as the proximal and distal axial ligands in ferric LmPP, respectively. However, the electronic properties of reduced wild-type LmPP are similar to those of known 5cHS His-ligated heme proteins at pH 8.8, indicating that the thiolate bond was broken upon reduction. Furthermore, the wild-type protein was only partially reduced at pH 4.0, but the E105L mutant was completely reduced to form a 5cHS ferrous heme. These results imply that the presence of an acidic residue near the distal site may prevent reduction of the heme iron at acidic pH.

show abstract

Section: ■ Discussionmentioning

confidence: 70%

Identification of Proximal and Distal Axial Ligands in Leishmania major Pseudoperoxidase

et al. 2013

View full text Add to dashboard Cite

show abstract

“…In making substitutions to His261 and His491, we observed that a His261Cys substitution in CcmF retained a high level of function relative to WT (see below) and that purified CcmF His261Cys (Figure A) contained ∼50% of the b -heme as WT (Figure B). Cysteine has been shown to function as a ligand in place of the natural histidine ligand in many heme proteins, including myoglobin, − cytochrome b 5 , and flavocytochrome b 2 . The UV/vis absorption spectrum of CcmF His261Cys has several distinct features in comparison to the spectrum of WT CcmF (compare parts C and D of Figure ).…”

Section: Resultsmentioning

confidence: 99%

Heme Ligand Identification and Redox Properties of the Cytochrome c Synthetase, CcmF

et al. 2011

View full text Add to dashboard Cite

Cytochrome c maturation in many bacteria, archaea, and plant mitochondria involves the integral membrane protein CcmF, which is thought to function as a cytochrome c synthetase by facilitating the final covalent attachment of heme to the apocytochrome c. We previously reported that the E. coli CcmF protein contains a b-type heme that is stably and stoichiometrically associated with the protein and is not the heme attached to apocytochrome c. Here, we show that mutation of either of two conserved transmembrane histidines (His261 or His491) impairs stoichiometric b-heme binding in CcmF and results in spectral perturbations in the remaining heme. Exogeneous imidazole is able to correct cytochrome c maturation for His261 and His491 substitutions with small side chains (Ala or Gly), suggesting that a “cavity” is formed in these CcmF mutants in which imidazole binds and acts as a functional ligand to the b-heme. The results of resonance Raman spectroscopy on wild-type CcmF are consistent with a hexacoordinate low spin b-heme with at least one endogeneous axial His ligand. Analysis of purified recombinant CcmF proteins from diverse prokaryotes reveals that the b-heme in CcmF is widely conserved. We have also determined the reduction potential of the CcmF b-heme (Em,7 = -147 mV). We discuss these results in the context of CcmF structure and functions as a heme reductase and cytochrome c synthetase.

show abstract

“…Fortunately, proximal ligand replacement via sitedirected mutagenesis has proven much more successful for myoglobin than for cytochrome P450s. The His-93 has been successfully replaced with cysteine in horse heart, 54 human, 43,55,56 and sperm whale myoglobins. 57 The most extensive functional studies on the H93C mutants were undertaken by Watanabe, Morishima, and co-workers with human myoglobin.…”

Section: The Role Of the Proximal Heme Iron Ligandmentioning

confidence: 99%

“…43,56 However, upon reduction to the ferrous state, both in the presence and in the absence of CO, none of the H93C mutants are able to retain their thiolate ligation. 43,54,56 The ratio of heterolysis/homolysis evaluated from the reaction with CHP (eqs 1-3) increases from 69% to 89% for the H93C single mutation, and further substitution of the distal histidine with a glycine or valine residue does not alter the ratio significantly. 56 Thus, our results have supported the hypothesis that the electron donation of the proximal ligand enhances heterolysis.…”

Section: The Role Of the Proximal Heme Iron Ligandmentioning

confidence: 99%

Investigations of the Roles of the Distal Heme Environment and the Proximal Heme Iron Ligand in Peroxide Activation by Heme Enzymes via Molecular Engineering of Myoglobin

et al. 2001

View full text Add to dashboard Cite

To pursue structure-function relationships of heme enzymes in the activation of peroxides, we have chosen to use myoglobin as the framework for our molecular engineering studies. Comparison of the crystal structures of myoglobin and peroxidases reveals differences in the arrangement of amino acid residues in heme active sites. On the basis of these structural differences and the reaction mechanisms of peroxidases, we have converted myoglobin into a peroxidase-like enzyme by alternation of the heme distal pocket via site-directed mutagenesis. The replacement of the proximal histidine with cysteine and the exogenous substituted imidazoles slightly accelerates the peroxide O-O bond cleavage due to the electron donor characteristics. However, we have not observed an enhancement in the activation of peroxide by the proximal mutant with tyrosine, the exogenous phenolate, and benzoate. A clear understanding of the absolute role of the proximal ligand remains elusive.

show abstract

Trans effects on cysteine ligation in the proximal HIS93CYS variant of horse heart myoglobin

Cited by 5 publications

References 0 publications

Identification of Proximal and Distal Axial Ligands in Leishmania major Pseudoperoxidase

Identification of Proximal and Distal Axial Ligands in Leishmania major Pseudoperoxidase

Heme Ligand Identification and Redox Properties of the Cytochrome c Synthetase, CcmF

Investigations of the Roles of the Distal Heme Environment and the Proximal Heme Iron Ligand in Peroxide Activation by Heme Enzymes via Molecular Engineering of Myoglobin

Contact Info

Product

Resources

About