MinireviewRedox regulation and reaction mechanism of human cystathionine-β-synthase: a PLP-dependent hemesensor protein☆
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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 forserine 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,
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This work was supported by a grant from the National Institutes of Health (HL58984).