Effects of the loss of the axial tyrosine ligand of the low‐spin heme of MauG on its physical properties and reactivity

Tarboush, Nafez Abu; Shin, Sooim; Geng, Jiafeng; Liu, Aimin; Davidson, Victor L.

doi:10.1016/j.febslet.2012.10.044

Cited by 16 publications

(16 citation statements)

References 28 publications

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“…Of these, HasA, PhuR, CcmE and HmuT are all involved in heme transfer. MauG, in contrast, appears to use the tyrosinate axial ligand to stabilize a high oxidation center in the mechanistic pathway of this protein (116). The His/Tyr ligand set is also known in hemoglobin variants such as Hb M Saskatoon (102) and has been created by site-directed mutagenesis of sperm whale Mb (HisE7Tyr) (103).…”

Section: Discussionmentioning

confidence: 99%

Heme Binding by Corynebacterium diphtheriae HmuT: Function and Heme Environment

et al. 2015

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The heme uptake pathway (hmu) of Corynebacterium diphtheriae utilizes multiple proteins to bind and transport heme into the cell. One of these proteins, HmuT, delivers heme to the ABC transporter HmuUV. In this study, the axial ligation of the heme in ferric HmuT is probed by examination of wild-type HmuT and a series of conserved heme pocket residue mutants, H136A, Y235A, and M292A. Characterization by UV-visible, resonance Raman, and magnetic circular dichroism spectroscopies indicate that H136 and Y235 are the axial ligands in ferric HmuT. Consistent with this assignment of axial ligands, ferric WT and H136A HmuT are difficult to reduce while Y235A reduces readily in the presence of dithionite. Raman frequencies of the FeCO distortions in WT, H136A, and Y235A HmuT–CO complexes provide further evidence for the axial ligand assignments. Additionally, the se frequencies provide insight into the nonbonding environment of the heme pocket. Ferrous Y235A and the Y235A–CO complex reveal that the imidazole of H136 exists in two forms, one neutral and one with imidazolate character, consistent with a hydrogen-bond acceptor on the H136 side of the heme. The ferric fluoride complex of Y235A reveals the presence of at least one hydrogen-bond donor on the Y235 side of the heme. Hemoglobin utilization assays showed that the axial Y235 ligand is required for heme uptake in HmuT.

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Section: Discussionmentioning

confidence: 99%

Heme Binding by Corynebacterium diphtheriae HmuT: Function and Heme Environment

et al. 2015

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“…Site-directed mutagenesis [17] and kinetic and thermodynamic analyses [21] showed that Trp 199 mediates a hole-hopping mechanism of ET that is required for MauG-dependent TTQ biosynthesis. Previous studies also demonstrated that a critical role of the bis-Fe IV intermediate of MauG was not only to provide a high-potential oxidant for modification of preMADH, but also to shorten the distance for the required long-range ET from the substrate to the nearest Fe IV haem [14,15].…”

Section: Introductionmentioning

confidence: 98%

“…These oxidations proceed via a high-valence bis-Fe IV intermediate of MauG [13] in which one haem is present as Fe IV = O with an axial ligand provided by a histidine residue and the other is present as Fe IV with histidine-tyrosine axial ligation and no exogenous bound ligand [8]. It has been shown by site-directed mutagenesis that the tyrosine ligation of the six-co-ordinate haem is critical for stabilization of the bis-Fe IV intermediate [14,15]. This redox state is generated and stabilized by MauG despite the fact that the two haem irons are separated by 21 Å (1 Å = 0.1 nm) (Figure 2A).…”

Section: Introductionmentioning

confidence: 99%

Mutation of Trp93 of MauG to tyrosine causes loss of bound Ca2+ and alters the kinetic mechanism of tryptophan tryptophylquinone cofactor biosynthesis

Shin¹,

Feng

Davidson³

2013

Biochemical Journal

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The dihaem enzyme MauG catalyses a six-electron oxidation required for post-translational modification of preMADH (precursor of methylamine dehydrogenase) to complete the biosynthesis of its TTQ (tryptophan tryptophylquinone) cofactor. Trp93 of MauG is positioned midway between its two haems, and in close proximity to a Ca2+ that is critical for MauG function. Mutation of Trp93 to tyrosine caused loss of bound Ca2+ and changes in spectral features similar to those observed after removal of Ca2+ from WT (wild-type) MauG. However, whereas Ca2+-depleted WT MauG is inactive, W93Y MauG exhibited TTQ biosynthesis activity. The rate of TTQ biosynthesis from preMADH was much lower than that of WT MauG and exhibited highly unusual kinetic behaviour. The steady-state reaction exhibited a long lag phase, the duration of which was dependent on the concentration of preMADH. The accumulation of reaction intermediates, including a diradical species of preMADH and quinol MADH (methylamine dehydrogenase), was detected during this pre-steady-state phase. In contrast, steady-state oxidation of quinol MADH to TTQ, the final step of TTQ biosynthesis, exhibited no lag phase. A kinetic model is presented to explain the long pre-steady-state phase of the reaction of W93Y MauG, and the role of this conserved tryptophan residue in MauG and related dihaem enzymes is discussed.

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“…These results are consistent with a mixed-valent state in which only one heme has been reduced. A mixed-valent state was not observed for WT MauG, or all but one of the many other variant MauG proteins which have been previously studied [7, 10, 17, 18, 22, 26]. The exception is E113Q MauG, which like T67A MauG could only be reduced by dithionite to a mixed-valent state which exhibited an absorbance spectrum very similar to that of T67A MauG [25].…”

Section: Resultsmentioning

confidence: 99%

A T67A mutation in the proximal pocket of the high-spin heme of MauG stabilizes formation of a mixed-valent FeII/FeIII state and enhances charge resonance stabilization of the bis-FeIV state

Shin

Feng

et al. 2015

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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The diheme enzyme MauG catalyzes a six-electron oxidation required for posttranslational modification of a precursor of methylamine dehydrogenase (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. One heme is low-spin with ligands provided by His205 and Tyr294, and the other is high-spin with a ligand provided by His35. The side chain methyl groups of Thr67 and Leu70 are positioned at a distance of 3.4 Å on either side of His35, maintaining a hydrophobic environment in the proximal pocket of the high-spin heme and restricting the movement of this ligand. Mutation of Thr67 to Ala in the proximal pocket of the high-spin heme prevented reduction of the low-spin heme by dithionite, yielding a mixed-valent state. The mutation also enhanced the stabilization of the charge-resonance-transition of the high-valent bis-FeIV state that is generated by addition of H2O2. The rates of electron transfer from TTQ biosynthetic intermediates to the high-valent form of T67A MauG were similar to that of wild-type MauG. These results are compared to those previously reported for mutation of residues in the distal pocket of the high-spin heme that also affected the redox properties and charge resonance transition stabilization of the high-valent state of the hemes. However, given the position of residue 67, the structure of the variant protein and the physical nature of the T67A mutation, the basis for the effects of the T67A mutation must be different from those of the mutations of the residues in the distal heme pocket.

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Effects of the loss of the axial tyrosine ligand of the low‐spin heme of MauG on its physical properties and reactivity

Cited by 16 publications

References 28 publications

Heme Binding by Corynebacterium diphtheriae HmuT: Function and Heme Environment

Heme Binding by Corynebacterium diphtheriae HmuT: Function and Heme Environment

Mutation of Trp93 of MauG to tyrosine causes loss of bound Ca2+ and alters the kinetic mechanism of tryptophan tryptophylquinone cofactor biosynthesis

A T67A mutation in the proximal pocket of the high-spin heme of MauG stabilizes formation of a mixed-valent FeII/FeIII state and enhances charge resonance stabilization of the bis-FeIV state

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