Mercaptosuccinate Dioxygenase, a Cysteine Dioxygenase Homologue, from Variovorax paradoxus Strain B4 Is the Key Enzyme of Mercaptosuccinate Degradation

Brandt, Ulrike; Schürmann, Marc; Steinbüchel, Alexander

doi:10.1074/jbc.m114.579730

Cited by 27 publications

(27 citation statements)

References 38 publications

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“…This common observation of higher IC 50 values observed for both cell types could be due to the low intracellular availability of the drug (owing to the hydrophilic nature of the molecule and hence poor transport) and/or due to metabolism leading to degradation of MSA. MSA dioxygenase, an enzyme that converts mercaptosuccinic acid to succinate is present in the bacterium Variovorax paradoxus (48,49). We, however, could not find homologues of the enzyme in E. coli or P. falciparum.…”

Section: Discussionmentioning

confidence: 49%

Biochemical characterization and essentiality of fumarate hydratase

Jayaraman¹,

Suryavanshi²,

Kalale

et al. 2018

Journal of Biological Chemistry

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Plasmodium falciparum (Pf), the causative agent of malaria, has an iron-sulfur cluster-containing class I fumarate hydratase (FH) that catalyzes the interconversion of fumarate to malate, a wellknown reaction in the tricarboxylic acid cycle. In humans, the same reaction is catalyzed by class II FH that has no sequence or structural homology with the class I enzyme from Plasmodium. Fumarate is generated in large quantities in the parasite as a byproduct of AMP synthesis and is converted to malate by FH and then used in the generation of the key metabolites oxaloacetate, aspartate, and pyruvate. Previous studies have identified the FH reaction as being essential to P. falciparum, but biochemical characterizations of PfFH that may provide leads for the development of specific inhibitors are lacking. Here, we report on the kinetic characterization of purified recombinant PfFH, functional complementation of fh deficiency in Escherichia coli, and mitochondrial localization in the parasite. We found that the substrate analog mercaptosuccinic acid is a potent PfFH inhibitor, with a K i value in the nanomolar range. The fh gene could not be knocked out in Plasmodium berghei when transfectants were introduced into BALB/c mice; however, fh knockout was successful when C57BL/6 mice were used as host, suggesting that the essentiality of the fh gene to the parasite was mouse strain dependent.Plasmodium falciparum (Pf), the causative agent of the most lethal form of malaria, during its intra-erythrocytic asexual stages, derives ATP primarily from glycolysis with low contribution from mitochondrial pathways (1, 2). The bulk of pyruvate formed is converted to lactic acid with a minor amount entering the tricarboxylic acid (TCA) cycle, the flux through which is upregulated in sexual stages (2). Key intermediates that anaplerotically feed into the TCA cycle are α-ketoglutarate derived from glutamate, oxaloacetate (OAA) from phosphoenolpyruvate, and fumarate from adenosine 5΄-monophosphate (AMP) synthesis. Synthesis of AMP in the parasite is solely from inosine-5΄-monophosphate (IMP) through a pathway involving the enzymes adenylosuccinate synthetase (ADSS) and adenylosuccinate lyase (ASL). The net reaction of ADSS and ASL involves consumption of GTP and aspartate and, generation of GDP, P i and fumarate. In the rapidly dividing parasite with an AT-rich genome and high energy requirements, leading to a high demand for adenine pools, one would expect a high flux of fumarate generation. The parasite does not secrete fumarate but instead, the carbon derived from this metabolite can be traced in malate, OAA, aspartate, pyruvate (through PEP) and lactate (3). The metabolic significance of this fumarate anaplerosis is still obscure. In this context, fumarate hydratase (FH, fumarase) the key enzyme to metabolize fumarate becomes an Fumarate hydratase (fumarase, E.C. 4.2.1.2) catalyzes the reversible conversion of fumarate to malate. The stereospecific reaction involves the anti-addition of a water molecule across the carbon-carbon...

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Section: Discussionmentioning

confidence: 49%

Biochemical characterization and essentiality of fumarate hydratase

Jayaraman¹,

Suryavanshi²,

Kalale

et al. 2018

Journal of Biological Chemistry

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“…Mammalian ADO, bacterial mercaptosuccinate dioxygenase, and bacterial 3MDO are only able to dioxygenate cysteamine, mercaptosuccinate, and 3-MPA, respectively (6,83,84). Although recent work (80) has shown mouse CDO is able to oxidize other substrates, such as L-penicillamine, the coupling between dioxygen consumption and product formation is Յ5%, suggesting that these substrates interact with the enzyme active site and undergo catalysis very differently from the authentic substrate.…”

Section: Discussionmentioning

confidence: 91%

The Cysteine Dioxygenase Homologue from Pseudomonas aeruginosa Is a 3-Mercaptopropionate Dioxygenase

Tchesnokov

Fellner

Siakkou

et al. 2015

Journal of Biological Chemistry

117

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Background: Thiol dioxygenation is catalyzed by enzymes specific for each substrate. Results: Kinetic, structural, and spectroscopic data describe an enzyme from P. aeruginosa that is a 3-mercaptopropionate dioxygenase with secondary cysteine dioxygenase activity. Conclusion: An arginine to glutamine switch and the absence of a cis-peptide bond correlate with substrate preference. Significance: Characterization of this enzyme deepens our understanding of substrate specificity in thiol dioxygenases.

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“…CDO is part of a larger class of enzymes, the thiol dioxygenases, that catalyze formation of the corresponding sulfinate from a range of different thiols. This group includes cysteamine dioxygenase, [1] 3-mercaptopropionate dioxygenase [2][3][4][5] and mercaptosuccinate dioxygenase, [6] which have been found to have a range of substrate specificities. CDO catalyzes the first step in oxidative cysteine breakdown in mammals.…”

Section: Introductionmentioning

confidence: 99%

Influence of cysteine 164 on active site structure in rat cysteine dioxygenase

et al. 2016

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Cysteine dioxygenase (CDO) is a non-heme mononuclear iron enzyme with unique structural features, namely an intramolecular thioether crosslink between cysteine 93 and tyrosine 157, and a disulfide between substrate L-cysteine and cysteine 164 in the entrance channel to the active site. We investigated how these posttranslational modifications affect catalysis through a kinetic, crystallographic and computational study. The enzyme kinetics of a C164S variant are identical to WT indicating that disulfide formation at C164 does not significantly impair access to the active site at physiological pH. However, at high pH the cysteine-tyrosine crosslink formation is enhanced in C164S. This supports the view that disulfide formation at position 164 can limit access to the active site. The C164S yielded crystal structures of unusual clarity in both resting state and with cysteine bound. Both show that the iron in the cysteine bound complex is a mixture of penta-and hexa-coordinate with a water molecule taking up the final site (60% occupancy), which is where dioxygen is believed to coordinate during turnover. The serine also displays stronger hydrogen bond interactions to a water bound to the amine of the substrate cysteine. However, the interactions between cysteine and iron appear unchanged. DFT calculations support this and show that WT and C164S have similar binding energies for the water molecule in the final site. This variant therefore provides evidence that WT also exists in this equilibrium between penta-and hexa-coordinate forms and presence of the sixth ligand does not strongly affect dioxygen binding.

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Mercaptosuccinate Dioxygenase, a Cysteine Dioxygenase Homologue, from Variovorax paradoxus Strain B4 Is the Key Enzyme of Mercaptosuccinate Degradation

Cited by 27 publications

References 38 publications

Biochemical characterization and essentiality of fumarate hydratase

Biochemical characterization and essentiality of fumarate hydratase

The Cysteine Dioxygenase Homologue from Pseudomonas aeruginosa Is a 3-Mercaptopropionate Dioxygenase

Influence of cysteine 164 on active site structure in rat cysteine dioxygenase

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