How Replacements of the 12 Conserved Histidines of Subunit I Affect Assembly, Cofactor Binding, and Enzymatic Activity of the Bradyrhizobium japonicum cbb 3-type Oxidase

Zufferey, Rachel; Arslan, Engin; Thöny‐Meyer, Linda; Hennecke, Hauke

doi:10.1074/jbc.273.11.6452

Cited by 39 publications

(37 citation statements)

References 40 publications

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“…According to mutation data, substitution of this histidine by alanine leads to fully assembled although inactive enzyme in Bradyrhizobium japonicum cytochrome cbb 3 . The Cu B center is also present despite the mutation, implying that the lack of activity is not due to lack of cofactor (20).…”

Section: Discussionmentioning

confidence: 99%

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Identification of a histidine-tyrosine cross-link in the active site of the cbb ₃ -type cytochrome c oxidase from Rhodobacter sphaeroides

Rauhamäki

Baumann

Soliymani

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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The heme-copper oxidases constitute a superfamily of terminal dioxygen-reducing enzymes located in the inner mitochondrial or in the bacterial cell membrane. The presence of a mechanistically important covalent bond between a histidine ligand of the copper ion (Cu B) in the active site and a generally conserved tyrosine residue nearby has been shown to exist in the canonical cytochrome c oxidases. However, according to sequence alignment studies, this critical tyrosine is missing from the subfamily of cbb3-type oxidases found in certain bacteria. Recently, homology modeling has suggested that a tyrosine residue located in a different helix might fulfill this role in these enzymes. Here, we show directly by methods of protein chemistry and mass spectrometry that there is indeed a covalent link between this tyrosine and the copper-ligating histidine. The identity of the cross-linked tyrosine was determined by showing that the cross-link is not formed when this residue is replaced by phenylalanine, even though structural integrity is maintained. These results suggest a universal functional importance of the histidine-tyrosine cross-link in the mechanism of O2 reduction by all heme-copper oxidases.mass spectrometry

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Section: Discussionmentioning

confidence: 99%

“…Although there is no three-dimensional structure available for the cbb 3 -type oxidases, it is likely that these histidines have the same roles also in these enzymes (20). The three histidine ligands of Cu B are all located in helices VI and VII (Fig.…”

Section: Discussionmentioning

confidence: 99%

Identification of a histidine-tyrosine cross-link in the active site of the cbb ₃ -type cytochrome c oxidase from Rhodobacter sphaeroides

Rauhamäki

Baumann

Soliymani

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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“…The presence of CcoQ in a purified complex has so far been demonstrated immunologically only in B. japonicum (52), and its function remains unclear. In-frame deletion mutants of ccoQ constructed in B. japonicum (23) and R. sphaeroides (53) have no apparent effect upon the assembly or activity of cytochrome cbb 3 oxidase, although the amount of cytochrome c appears to be somewhat decreased in the ⌬ccoQ mutant of B. japonicum (23).…”

Section: Discussionmentioning

confidence: 99%

Molecular and Spectroscopic Analysis of the Cytochromecbb3 Oxidase from Pseudomonas stutzeri

Pitcher¹,

Cheesman²,

Watmough³

2002

Journal of Biological Chemistry

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Cytochrome cbb 3 oxidase, a member of the heme-copper oxidase superfamily, is characterized by its high affinity for oxygen while retaining the ability to pump protons. These attributes are central to its proposed role in the microaerobic metabolism of proteobacteria. We have completed the first detailed spectroscopic characterization of a cytochrome cbb 3 oxidase, the enzyme purified from Pseudomonas stutzeri. A combination of UVvisible and magnetic CD spectroscopies clearly identified four low-spin hemes and the high-spin heme of the active site. This heme complement is in good agreement with our analysis of the primary sequence of the ccoNOPQ operon and biochemical analysis of the complex. Near-IR magnetic CD spectroscopy revealed the unexpected presence of a low-spin bishistidine-coordinated c-type heme in the complex. This was shown to be one of two c-type hemes in the CcoP subunit by separately expressing the subunit in Escherichia coli. Separate expression of CcoP also allowed us to unambiguously assign each of the signals associated with low-spin ferric hemes present in the X-band EPR spectrum of the oxidized enzyme. This work both underpins future mechanistic studies on this distinctive class of bacterial oxidases and raises questions concerning the role of CcoP in electron delivery to the catalytic subunit.

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“…Cell Fractionation and Analysis. Whole-cell protein analysis, isolation of periplasmic and membrane fractions, determination of protein concentration, and heme staining after SDS͞PAGE were performed as described previously (7,(17)(18)(19). Western blot detection was performed with CcmE-specific antiserum (7) and with mAbs directed against a tetra-His epitope (Qiagen).…”

Section: Methodsmentioning

confidence: 99%

Heme transfer to the heme chaperone CcmE during cytochrome c maturation requires the CcmC protein, which may function independently of the ABC-transporter CcmAB

Schulz

Fabianek²,

Pellicioli³

et al. 1999

Proc. Natl. Acad. Sci. U.S.A.

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Cytochrome c maturation in Escherichia coli requires the ccm operon, which encodes eight membrane proteins (CcmABCDEFGH). CcmE is a periplasmic heme chaperone that binds heme covalently and transfers it onto apocytochrome c in the presence of CcmF, CcmG, and CcmH. In this work we addressed the functions of the ccmABCD gene products with respect to holo-CcmE formation and the subsequent ligation of heme to apocytochrome c. In the absence of the ccmABCD genes, heme is not bound to CcmE. We report that CcmC is functionally uncoupled from the ABC transporter subunits CcmA and CcmB, because it is the only Ccm protein that is strictly required for heme transfer and attachment to CcmE. Site-directed mutagenesis of conserved histidines inactivates the CcmC protein, which is in agreement with the hypothesis that this protein interacts directly with heme. We also present evidence that questions the role of CcmAB as a heme exporter; yet, the transported substrate remains unknown. CcmD was found to be involved in stabilizing the heme chaperone CcmE in the membrane. We propose a heme-trafficking pathway as part of a substantially revised model for cytochrome c maturation in E. coli.

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How Replacements of the 12 Conserved Histidines of Subunit I Affect Assembly, Cofactor Binding, and Enzymatic Activity of the Bradyrhizobium japonicum cbb 3-type Oxidase

Cited by 39 publications

References 40 publications

Identification of a histidine-tyrosine cross-link in the active site of the cbb ₃ -type cytochrome c oxidase from Rhodobacter sphaeroides

Identification of a histidine-tyrosine cross-link in the active site of the cbb ₃ -type cytochrome c oxidase from Rhodobacter sphaeroides

Molecular and Spectroscopic Analysis of the Cytochromecbb3 Oxidase from Pseudomonas stutzeri

Heme transfer to the heme chaperone CcmE during cytochrome c maturation requires the CcmC protein, which may function independently of the ABC-transporter CcmAB

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How Replacements of the 12 Conserved Histidines of Subunit I Affect Assembly, Cofactor Binding, and Enzymatic Activity of the Bradyrhizobium japonicum cbb 3-type Oxidase

Cited by 39 publications

References 40 publications

Identification of a histidine-tyrosine cross-link in the active site of the cbb 3 -type cytochrome c oxidase from Rhodobacter sphaeroides

Identification of a histidine-tyrosine cross-link in the active site of the cbb 3 -type cytochrome c oxidase from Rhodobacter sphaeroides

Molecular and Spectroscopic Analysis of the Cytochromecbb3 Oxidase from Pseudomonas stutzeri

Heme transfer to the heme chaperone CcmE during cytochrome c maturation requires the CcmC protein, which may function independently of the ABC-transporter CcmAB

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Product

Resources

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Identification of a histidine-tyrosine cross-link in the active site of the cbb ₃ -type cytochrome c oxidase from Rhodobacter sphaeroides

Identification of a histidine-tyrosine cross-link in the active site of the cbb ₃ -type cytochrome c oxidase from Rhodobacter sphaeroides