Flavodoxins reactivity

The oxygen reactivity of flavohydroquinone bound to apoflavoprotein dehydrogenases can vary considerably from fast (flavodoxins), moderate (xanthine oxidase) to nil (succinate dehydrogenase) Most, but not all, flavoprotein dehydrogenases contain one or more types of metal prosthetic groups, e.g. xanthine oxidase contains also Fe and Mo. Since these metal ions are involved in electron flux, their possible participation in the reaction with O2 cannot be excluded. Much evidence, however, indicates that the flavin is involved in the one-electron reduction of Oj, as shown in Equation (5). 

MetH catalyzes the methylation of the bound and reduced cob(I)alamin cofactor by (N -protonated) N -methyltetrahydrofolate to give enzyme-bound methylcobalamin (3) in a base-off/His-on form (see later) [125,153-155]. The methyl-Co(III)corrinoid is demethylated by homocysteine, whose sulfur is activated and deprotonated due to the coordination to a zinc ion (held by three cysteine residues) of the homocysteine binding domain [164] (see Fig. 15). The two methyl-transfer reactions occur in a sequential mechanism [124,125,153,154]. Intermittently, the bound Cob(I)alamin (40 ) is oxidized to enzymatically inactive cob(II)alamin (23) and requires reactivation by reductive methylation with SAM and a flavodoxin as a reducing agent [125,153-155,165]. 

The primary reaction catalyzed by methionine synthase converts homocysteine (Hey) and methyltetrahydrofolate (CH3H4folate) to methionine and tetrahydrofolate (Figure 2). Occasional oxidation of the reactive cob(I)alamin intermediate produces an inactive cob(II)alamin enzyme, which is reactivated by a reductive methylation that uses S-adenosylmethionine (AdoMet) as the methyl donor and flavodoxin or a flavodoxin-like domain as an electron donor. Thus methionine synthase supports three distinct methyl transfer reactions each involving the cobalamin cofactor. 

Figure 2, Catalysis and reactivation of methionine synthase. Methionine for,motion occurs via two half reactions in which cobalamin serves as the intermediate methyl carrier. Reactivation is depicted in the right-hand portion of the diagram. An electron donor and AdoMet convert the inactive cob(II)alamin form of the enzyme to methylcob(III)alamin. In E. colU flavodoxin serves as the reductant for this priming reaction (7).

The simplest scheme for reactivation of the cob(II)alamin form of E. coli MetH involves two steps reduction to cob(I)alamin by flavodoxin and subsequent methylation by AdoMet (Figure 1). The first step presents a thermodynamic problem, since flavodoxin potentials, even those for the semiquinone/hydroquinone equilibrium, are more positive than the MetH cob(II)/cob(I) potentials. Over a decade ago Banerjee et aL (39) showed convincingly that reduction can proceed even at rather high potential when driven by coupling with the favorable AG of transfer from AdoMet. Indeed these thermodynamics provide the rationale for deploying AdoMet in reactivation. 

Recent investigations of the in vitro reactivation of cob(II) MetH by flavodoxin hydroquinone (40) have provided a more complete description of the reaction. Initially, only a small fraction of the enzyme is in a conformation suitable for reaction with flavodoxin, and this fraction is rapidly reduced to cob(I)alamin. For the remainder of the enzyme, reduction is preceded by a very slow step involving dissociation of the histidine 759 ligand to form a 4-coordinate cob(II)alamin species. The unligated (base-off) intermediate can then be reduced by flavodoxin, and cob(I)alamin is methylated and finally religated to His. Rate constants for formation and decay of intermediates were obtained from reactions with the mutant Asp757Glu, which is mostly 4-coordinate in the cob(II)alamin state. 

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