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A Trp199Glu MauG variant reveals a role for Trp199 interactions with pre-methylamine dehydrogenase during tryptophan tryptophylquinone biosynthesis.

ABSTRACT: MauG catalyzes posttranslational modifications of a methylamine dehydrogenase precursor (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. Trp199 is present at the site of interaction between MauG and preMADH and is critical to this process as it mediates hole hopping during the inter-protein electron transfer that is required for catalysis. Trp199 was converted to Glu and the structure and reactivity of the W199E/preMADH complex were characterized. The results reveal that the nature of residue 199 is also important for productive complex formation between preMADH and MauG.

SUBMITTER: Abu Tarboush N

PROVIDER: S-EPMC3747028 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature

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A Trp199Glu MauG variant reveals a role for Trp199 interactions with pre-methylamine dehydrogenase during tryptophan tryptophylquinone biosynthesis.

Abu Tarboush Nafez N Jensen Lyndal M R LM Wilmot Carrie M CM Davidson Victor L VL

FEBS letters 20130510 12

MauG catalyzes posttranslational modifications of a methylamine dehydrogenase precursor (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. Trp199 is present at the site of interaction between MauG and preMADH and is critical to this process as it mediates hole hopping during the inter-protein electron transfer that is required for catalysis. Trp199 was converted to Glu and the structure and reactivity of the W199E/preMADH complex were chara ...[more]

PMID: 23669364

Similar Datasets

Long-range electron transfer reactions between hemes of MauG and different forms of tryptophan tryptophylquinone of methylamine dehydrogenase.

Project description:The diheme enzyme MauG catalyzes the post-translational modification of a precursor protein of methylamine dehydrogenase (preMADH) to complete the biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. This six-electron oxidation of preMADH requires long-range electron transfer (ET) as the structure of the MauG-preMADH complex reveals that the shortest distance between the modified residues of preMADH and the nearest heme of MauG is 14.0 A [Jensen, L. M. R., Sanishvili, R., Davidson, V. L., and Wilmot, C. M. (2010) Science 327, 1392-1394]. The kinetics of two ET reactions between MADH and MauG have been analyzed. Interprotein ET from quinol MADH to the high-valent bis-Fe(IV) form of MauG exhibits a K(d) of 11.2 microM and a rate constant of 20 s(-1). ET from diferrous MauG to oxidized TTQ of MADH exhibits a K(d) of 10.1 microM and a rate constant of 0.07 s(-1). These similar K(d) values are much greater than that for the MauG-preMADH complex, indicating that the extent of TTQ maturity rather than its redox state influences complex formation. The difference in rate constants is consistent with a larger driving force for the faster reaction. Analysis of the structure of the MauG-preMADH complex in the context of ET theory and these results suggests that direct electron tunneling between the residues that form TTQ and the five-coordinate oxygen-binding heme is not possible, and that ET requires electron hopping via the six-coordinate heme.

| S-EPMC2913433 | biostudies-literature

MauG, a diheme enzyme that catalyzes tryptophan tryptophylquinone biosynthesis by remote catalysis.

Project description:MauG contains two c-type hemes with atypical physical and catalytic properties. While most c-type cytochromes function simply as electron transfer mediators, MauG catalyzes the completion of tryptophan tryptophylquinone (TTQ)(1) biosynthesis within a precursor protein of methylamine dehydrogenase. This posttranslational modification is a six-electron oxidation that requires crosslinking of two Trp residues, oxygenation of a Trp residue and oxidation of the resulting quinol to TTQ. These reactions proceed via a bis-Fe(IV) state in which one heme is present as Fe(IV)O and the other is Fe(IV) with axial heme ligands provided by His and Tyr side chains. Catalysis does not involve direct contact between the protein substrate and either heme of MauG. Instead it is accomplished by remote catalysis using a hole hopping mechanism of electron transfer in which Trp residues of MauG are reversibly oxidized. In this process, long range electron transfer is coupled to the radical mediated chemical reactions that are required for TTQ biosynthesis.

| S-EPMC3946517 | biostudies-literature

The tightly bound calcium of MauG is required for tryptophan tryptophylquinone cofactor biosynthesis.

Project description: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. The crystal structure of the MauG-preMADH complex revealed the presence of a Ca(2+) in proximity to the two hemes [Jensen, L. M. R., Sanishvili, R., Davidson, V. L., and Wilmot, C. M. (2010) Science 327, 1392-1394]. This Ca(2+) did not readily dissociate; however, after extensive treatment with EGTA or EDTA MauG was no longer able to catalyze TTQ biosynthesis and exhibited altered absorption and resonance Raman spectra. The changes in spectral features are consistent with Ca(2+)-dependent changes in heme spin state and conformation. Addition of H(2)O(2) to the Ca(2+)-depleted MauG did not yield spectral changes characteristic of formation of the bis-Fe(IV) state which is stabilized in native MauG. After addition of Ca(2+) to the Ca(2+)-depleted MauG, full TTQ biosynthesis activity and reactivity toward H(2)O(2) were restored, and the spectral properties returned to those of native MauG. Kinetic and equilibrium studies of Ca(2+) binding to Ca(2+)-depleted MauG indicated a two-step mechanism. Ca(2+) initially reversibly binds to Ca(2+)-depleted MauG (K(d) = 22.4 μM) and is followed by a relatively slow (k = 1.4 × 10(-3) s(-1)) but highly favorable (K(eq) = 4.2) conformational change, yielding an equilibrium dissociation constant K(d,eq) value of 5.3 μM. The circular dichroism spectra of native and Ca(2+)-depleted MauG were essentially the same, consistent with Ca(2+)-induced conformational changes involving domain or loop movements rather than general unfolding or alteration of secondary structure. These results are discussed in the context of the structures of MauG and heme-containing peroxidases.

| S-EPMC3061978 | biostudies-literature

In crystallo posttranslational modification within a MauG/pre-methylamine dehydrogenase complex.

Project description:MauG is a diheme enzyme responsible for the posttranslational modification of two tryptophan residues to form the tryptophan tryptophylquinone (TTQ) cofactor of methylamine dehydrogenase (MADH). MauG converts preMADH, containing monohydroxylated betaTrp57, to fully functional MADH by catalyzing the insertion of a second oxygen atom into the indole ring and covalently linking betaTrp57 to betaTrp108. We have solved the x-ray crystal structure of MauG complexed with preMADH to 2.1 angstroms. The c-type heme irons and the nascent TTQ site are separated by long distances over which electron transfer must occur to achieve catalysis. In addition, one of the hemes has an atypical His-Tyr axial ligation. The crystalline protein complex is catalytically competent; upon addition of hydrogen peroxide, MauG-dependent TTQ synthesis occurs.

| S-EPMC2878131 | biostudies-literature

Mutagenesis of tryptophan199 suggests that hopping is required for MauG-dependent tryptophan tryptophylquinone biosynthesis.

Project description:The diheme enzyme MauG catalyzes the posttranslational modification of the precursor protein of methylamine dehydrogenase (preMADH) to complete biosynthesis of its protein-derived tryptophan tryptophylquinone (TTQ) cofactor. Catalysis proceeds through a high valent bis-Fe(IV) redox state and requires long-range electron transfer (ET), as the distance between the modified residues of preMADH and the nearest heme iron of MauG is 19.4 Å. Trp199 of MauG resides at the MauG-preMADH interface, positioned midway between the residues that are modified and the nearest heme. W199F and W199K mutations did not affect the spectroscopic and redox properties of MauG, or its ability to stabilize the bis-Fe(IV) state. Crystal structures of complexes of W199F/K MauG with preMADH showed no significant perturbation of the MauG-preMADH structure or protein interface. However, neither MauG variant was able to synthesize TTQ from preMADH. In contrast, an ET reaction from diferrous MauG to quinone MADH, which does not require the bis-Fe(IV) intermediate, was minimally affected by the W199F/K mutations. W199F/K MauGs were able to oxidize quinol MADH to form TTQ, the putative final two-electron oxidation of the biosynthetic process, but with k(cat)/K(m) values approximately 10% that of wild-type MauG. The differential effects of the W199F/K mutations on these three different reactions are explained by a critical role for Trp199 in mediating multistep hopping from preMADH to bis-Fe(IV) MauG during the long-range ET that is required for TTQ biosynthesis.

| S-EPMC3193188 | biostudies-literature

Diradical intermediate within the context of tryptophan tryptophylquinone biosynthesis.

Project description:Despite the importance of tryptophan (Trp) radicals in biology, very few radicals have been trapped and characterized in a physiologically meaningful context. Here we demonstrate that the diheme enzyme MauG uses Trp radical chemistry to catalyze formation of a Trp-derived tryptophan tryptophylquinone cofactor on its substrate protein, premethylamine dehydrogenase. The unusual six-electron oxidation that results in tryptophan tryptophylquinone formation occurs in three discrete two-electron catalytic steps. Here the exact order of these oxidation steps in the processive six-electron biosynthetic reaction is determined, and reaction intermediates are structurally characterized. The intermediates observed in crystal structures are also verified in solution using mass spectrometry. Furthermore, an unprecedented Trp-derived diradical species on premethylamine dehydrogenase, which is an intermediate in the first two-electron step, is characterized using high-frequency and -field electron paramagnetic resonance spectroscopy and UV-visible absorbance spectroscopy. This work defines a unique mechanism for radical-mediated catalysis of a protein substrate, and has broad implications in the areas of applied biocatalysis and understanding of oxidative protein modification during oxidative stress.

| S-EPMC3607037 | biostudies-literature

Tryptophan tryptophylquinone biosynthesis: a radical approach to posttranslational modification.

Project description:Protein-derived cofactors are formed by irreversible covalent posttranslational modification of amino acid residues. An example is tryptophan tryptophylquinone (TTQ) found in the enzyme methylamine dehydrogenase (MADH). TTQ biosynthesis requires the cross-linking of the indole rings of two Trp residues and the insertion of two oxygen atoms onto adjacent carbons of one of the indole rings. The diheme enzyme MauG catalyzes the completion of TTQ within a precursor protein of MADH. The preMADH substrate contains a single hydroxyl group on one of the tryptophans and no crosslink. MauG catalyzes a six-electron oxidation that completes TTQ assembly and generates fully active MADH. These oxidation reactions proceed via a high valent bis-Fe(IV) state in which one heme is present as Fe(IV)=O and the other is Fe(IV) with both axial heme ligands provided by amino acid side chains. The crystal structure of MauG in complex with preMADH revealed that catalysis does not involve direct contact between the hemes of MauG and the protein substrate. Rather it is accomplished through long-range electron transfer, which presumably generates radical intermediates. Kinetic, spectrophotometric, and site-directed mutagenesis studies are beginning to elucidate how the MauG protein controls the reactivity of the hemes and mediates the long range electron/radical transfer required for catalysis. This article is part of a Special Issue entitled: Radical SAM enzymes and Radical Enzymology.

| S-EPMC3432176 | biostudies-literature

Posttranslational biosynthesis of the protein-derived cofactor tryptophan tryptophylquinone.

Project description:Methylamine dehydrogenase (MADH) catalyzes the oxidative deamination of methylamine to formaldehyde and ammonia. Tryptophan tryptophylquinone (TTQ) is the protein-derived cofactor of MADH required for this catalytic activity. TTQ is biosynthesized through the posttranslational modification of two tryptophan residues within MADH, during which the indole rings of two tryptophan side chains are cross-linked and two oxygen atoms are inserted into one of the indole rings. MauG is a c-type diheme enzyme that catalyzes the final three reactions in TTQ formation. In total, this is a six-electron oxidation process requiring three cycles of MauG-dependent two-electron oxidation events using either H2O2 or O2. The MauG redox form responsible for the catalytic activity is an unprecedented bis-Fe(IV) species. The amino acids of MADH that are modified are ≈ 40 Å from the site where MauG binds oxygen, and the reaction proceeds by a hole hopping electron transfer mechanism. This review addresses these highly unusual aspects of the long-range catalytic reaction mediated by MauG.

| S-EPMC4082410 | biostudies-literature

Crystal structures of CO and NO adducts of MauG in complex with pre-methylamine dehydrogenase: implications for the mechanism of dioxygen activation.

Project description:MauG is a diheme enzyme responsible for the post-translational formation of the catalytic tryptophan tryptophylquinone (TTQ) cofactor in methylamine dehydrogenase (MADH). MauG can utilize hydrogen peroxide, or molecular oxygen and reducing equivalents, to complete this reaction via a catalytic bis-Fe(IV) intermediate. Crystal structures of diferrous, Fe(II)-CO, and Fe(II)-NO forms of MauG in complex with its preMADH substrate have been determined and compared to one another as well as to the structure of the resting diferric MauG-preMADH complex. CO and NO each bind exclusively to the 5-coordinate high-spin heme with no change in ligation of the 6-coordinate low-spin heme. These structures reveal likely roles for amino acid residues in the distal pocket of the high-spin heme in oxygen binding and activation. Glu113 is implicated in the protonation of heme-bound diatomic oxygen intermediates in promoting cleavage of the O-O bond. Pro107 is shown to change conformation on the binding of each ligand and may play a steric role in oxygen activation by positioning the distal oxygen near Glu113. Gln103 is in a position to provide a hydrogen bond to the Fe(IV)?O moiety that may account for the unusual stability of this species in MauG.

| S-EPMC3071454 | biostudies-literature

Characterization of electron tunneling and hole hopping reactions between different forms of MauG and methylamine dehydrogenase within a natural protein complex.

Project description:Respiration, photosynthesis, and metabolism require the transfer of electrons through and between proteins over relatively long distances. It is critical that this electron transfer (ET) occur with specificity to avoid cellular damage, and at a rate that is sufficient to support the biological activity. A multistep hole hopping mechanism could, in principle, enhance the efficiency of long-range ET through proteins as it does in organic semiconductors. To explore this possibility, two different ET reactions that occur over the same distance within the protein complex of the diheme enzyme MauG and different forms of methylamine dehydrogenase (MADH) were subjected to kinetic and thermodynamic analysis. An ET mechanism of single-step direct electron tunneling from diferrous MauG to the quinone form of MADH is consistent with the data. In contrast, the biosynthetic ET from preMADH, which contains incompletely synthesized tryptophan tryptophylquinone, to the bis-Fe(IV) form of MauG is best described by a two-step hole hopping mechanism. Experimentally determined ET distances matched the distances determined from the crystal structure that would be expected for single-step tunneling and multistep hopping. Experimentally determined relative values of electronic coupling (H(AB)) for the two reactions correlated well with the relative H(AB) values predicted from computational analysis of the structure. The rate of the hopping-mediated ET reaction is also 10-fold greater than that of the single-step tunneling reaction despite a smaller overall driving force for the hopping-mediated ET reaction. These data provide insight into how the intervening protein matrix and redox potentials of the electron donor and acceptor determine whether the ET reaction proceeds via single-step tunneling or multistep hopping.

| S-EPMC3490227 | biostudies-literature

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