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Redox regulation and reaction mechanism of human cystathionine-β-synthase: a PLP-dependent hemesensor protein

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Abstract

Cystathionine β-synthase in mammals lies at a pivotal crossroad in methionine metabolism directing flux toward cysteine synthesis and catabolism. The enzyme exhibits a modular organization and complex regulation. It catalyzes the β-replacement of the hydroxyl group of serine with the thiolate of homocysteine and is unique in being the only known pyridoxal phosphate-dependent enzyme that also contains heme b as a cofactor. The heme functions as a sensor and modulates enzyme activity in response to redox change and to CO binding. Mutations in this enzyme are the single most common cause of hereditary hyperhomocysteinemia. Elucidation of the crystal structure of a truncated and highly active form of the human enzyme containing the heme- and pyridoxal phosphate binding domains has afforded a structural perspective on mechanistic and mutation analysis studies. The C-terminal regulatory domain containing two CBS motifs exerts intrasteric regulation and binds the allosteric activator, S-adenosylmethionine. Studies with mammalian cells in culture as well as with animal models have unraveled multiple layers of regulation of cystathionine β-synthase in response to redox perturbations and reveal the important role of this enzyme in glutathione-dependent redox homestasis. This review discusses the recent advances in our understanding of the structure, mechanism, and regulation of cystathionine β-synthase from the perspective of its physiological function, focusing on the clinically relevant human enzyme.

Section snippets

Cystathionine β-synthase is a modular protein

The human enzyme comprises 551 amino acids with a subunit molecular weight of ∼63 kDa [7]. It is a tetramer, which is prone to aggregation, and binds one heme and one PLP per subunit (Table 1). AdoMet functions as a V-type allosteric activator, binds stoichiometrically to each subunit [8], and increases enzyme activity ∼2- to 3-fold [8], [9], [10].

Cystathionine β-synthase displays a modular organization [8], [9], [10] in which an N-terminal heme domain is followed by a catalytic core that houses

Spectroscopic characteristics of the heme in cystathionine β-synthase

The heme in cystathionine β-synthase is spectroscopically distinct. A catalytic role for the heme in cystathionine β-synthase was excluded by spectroscopic studies that revealed that it was distant from the PLP, the site of β-replacement chemistry [24], was confirmed by the crystal structures [20], [21], and is consistent with its absence from the yeast enzyme, which catalyzes the same overall reaction, albeit without a second cofactor [14]. The presence of the heme is therefore enigmatic and

The reaction mechanism of cystathionine β-synthase

A constellation of conserved amino acids that interact with the cofactor have been described for the fold II family of PLP enzymes and are also found in cystathionine β-synthase (Fig. 5). The epsilon nitrogen of K119 in human cystathionine β-synthase extends from the back wall of the PLP binding pocket and forms a Schiff base with the aldehyde of PLP. S349 is positioned close to the pyridinium nitrogen of the cofactor, a position that is generally occupied by a polar residue (serine or

Alternative substrates and H2S formation

There have been a limited number of studies investigating the tolerance of cystathionine β-synthase to alternative substrates, some of which are listed in Eq. (2). β-Chloroalanine and threonine can substitute forX-CH2CH(NH2)COOH+YHXH+YCH2CH(NH2)COOHSerineHomocysteineCysteineH2SThreonineCH3CH2SHβ-ChloroalanineHomoserineAllothreonineCysteineserine in rat cystathionineβ-synthase and generate cystathionine and 3-methylcystathionine, respectively [42]. The relative efficiency of these

Intrasteric and allosteric regulation

The C-terminal domain of cystathionine β-synthase imparts interesting properties to the enzyme and regulates its activity via both intrasteric and allosteric effects (Fig. 7) and is important for maintaining the tetrameric state of the protein. Unfortunately, neither the structure of this domain nor of the full-length enzyme is available to provide a structural perspective on the conformational changes that the regulatory domain is predicted to undergo. A conserved protein folding motif, the

Redox regulation

Curiously, a subset of cystathionine β-synthases has two redox active switches that could, in principle, modulate enzyme activity. A heme cofactor and a CXXC oxidoreductase motif are found in the mammalian enzymes and are predicted, based on sequence homology, to exist in other organisms (Fig. 4). Under in vitro conditions, it is easier to test the influence of the redox changes in the heme than the CXXC motif on enzyme activity since the substrate, homocysteine, reduces disulfides, if present,

References (69)

  • S. Taoka et al.

    Evidence for heme-mediated redox regulation of human cystathionine β-synthase activity

    J. Biol. Chem.

    (1998)
  • S. Taoka et al.

    Mercuric chloride-induced spin or ligation state changes in ferric or ferrous human cystathionine b-synthase inhibit enzyme activity

    J. Inorg. Bioc.

    (2001)
  • M. Puranik et al.

    Dynamics of carbon monoxide binding to CooA

    J. Biol. Chem.

    (2004)
  • S. Taoka et al.

    Activity of human cystathionine beta synthase is regulated by CO and NO: possible role for the hemeprotein in CO sensing

    J. Inorg. Bioc.

    (2001)
  • S. Vadon-Le Goff et al.

    Coordination chemistry of the heme in cystathionine beta-synthase: formation of iron(II)-isonitrile complexes

    Biochem. Biophys. Res. Commun.

    (2001)
  • J. Oliveriusova et al.

    Deletion mutagenesis of human cystathionine beta-synthase. Impact on activity, oligomeric status, and S-adenosylmethionine regulation

    J. Biol. Chem.

    (2002)
  • K.H. Jhee et al.

    Mutation of an active site residue of tryptophan synthase (beta-serine 377) alters cofactor chemistry

    J. Biol. Chem.

    (1998)
  • S. Taoka et al.

    Stopped-flow kinetic analysis of the reaction catalyzed by the full length yeast cystathionine beta synthase

    J. Biol. Chem.

    (2002)
  • E. Borcsok et al.

    Mechanism of action of cystathionine synthase

    Arch. Biochem. Biophys.

    (1982)
  • M.V. Martinov et al.

    A substrate switch: A new mode of regulation in the methionine metabolic pathway

    J. Theor. Biol.

    (2000)
  • J.D. Finkelstein et al.

    Methionine metabolism in mammals. Distribution of homocysteine between competing pathways

    J. Biol. Chem.

    (1984)
  • W.M. Rabeh et al.

    Structure and mechanism of O-acetylserine sulfhydrylase

    J. Biol. Chem.

    (2004)
  • P. Burkhard et al.

    Ligand binding induces a large conformational change in O-acetylserine sulfhydrylase from Salmonella typhimurium

    J Mol Biol

    (1999)
  • S. Bruno et al.

    Functional properties of the active core of human cystathionine beta-synthase crystals

    J. Biol. Chem.

    (2001)
  • K. Eto et al.

    A novel enhancing mechanism for hydrogen sulfide-producing activity of cystathionine beta-synthase

    J. Biol. Chem.

    (2002)
  • M.D. Sintchak et al.

    Structure and mechanism of inosine monophosphate dehydrogenase in complex with the immunosuppressant mycophenolic acid

    Cell

    (1996)
  • O. Kabil et al.

    Deletion of the regulatory domain in the pyridoxal phosphate-dependent heme protein cystathionine beta-synthase alleviates the defect observed in a catalytic site mutant

    J. Biol. Chem.

    (1999)
  • M. Janosik et al.

    Impaired heme binding and aggregation of mutant cystathionine beta-synthase subunits in homocystinuria

    Am. J. Hum. Genet.

    (2001)
  • Zou et al.

    Tumor necrosis factor-a-induced targeted proteolysis of cystathionine beta-synthase modulates redox homeostasis

    J. Biol. Chem.

    (2003)
  • A. Meister

    Glutathione metabolism

    Methods Enzymol.

    (1995)
  • P.W. Beatty et al.

    Involvement of the cystathionine pathway in the biosynthesis of glutathione by isolated rat hepatocytes

    Arch. Biochem. Biophys.

    (1980)
  • M.H. Stipanuk

    Sulfur amino acid metabolism: pathways for production and removal of homocysteine and cysteine

    Annu. Rev. Nutr.

    (2004)
  • S.H. Mudd et al.

    The Metabolic and Molecular Basis of Inherited Diseases

    (1995)
  • J.P. Kraus et al.

    Cystathionine beta-synthase mutations in homocystinuria

    Hum. Mutat.

    (1999)
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    This work was supported by a grant from the National Institutes of Health (HL58984).

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