Abstract
The ferredoxin-dependent sulfite reductase from maize was treated, in separate experiments, with three different covalent modifiers of specific amino acid side chains. Treatment with the tryptophan-modifying reagent, N-bromosuccinimide (NBS), resulted in a loss of enzymatic activity with both the physiological donor for the enzyme, reduced ferredoxin, and with reduced methyl viologen, a non-physiological electron donor. Formation of the 1:1 ferredoxin/sulfite reductase complex prior to treating the enzyme with NBS completely protected the enzyme against the loss of both activities. Neither the secondary structure, nor the oxidation-reduction midpoint potential (E m) values of the siroheme and [4Fe–4S] cluster prosthetic groups of sulfite reductase, nor the binding affinity of the enzyme for ferredoxin were affected by NBS treatment. Treatment of sulfite reductase with the lysine-modifying reagent, N-acetylsuccinimide, inhibited the ferredoxin-linked activity of the enzyme without inhibiting the methyl viologen-linked activity. Complex formation with ferredoxin protects the enzyme against the inhibition of ferredoxin-linked activity produced by treatment with N-acetylsuccinimide. Treatment of sulfite reductase with N-acetylsuccinimide also decreased the binding affinity of the enzyme for ferredoxin. Treatment of sulfite reductase with the arginine-modifying reagent, phenylglyoxal, inhibited both the ferredoxin-linked and methyl viologen-linked activities of the enzyme but had a significantly greater effect on the ferredoxin-dependent activity than on the reduced methyl viologen-linked activity. The effects of these three inhibitory treatments are consistent with a possible role for a tryptophan residue the catalytic mechanism of sulfite reductase and for lysine and arginine residues at the ferredoxin-binding site of the enzyme.
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Abbreviations
- CD:
-
circular dichroism
- E h :
-
ambient oxidation-reduction potential
- E m :
-
oxidation-reduc-tion midpoint potential
- FNR:
-
ferredoxin:NADP+ oxidoreductase
- NBS:
-
N-bromosuccinimide
- Tris:
-
tris (hydroxymethyl)amino-methane
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Hirasawa, M., Nakayama, M., Kim, SK. et al. Chemical modification studies of tryptophan, arginine and lysine residues in maize chloroplast ferredoxin:sulfite oxidoreductase. Photosynth Res 86, 325–336 (2005). https://doi.org/10.1007/s11120-005-6966-y
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DOI: https://doi.org/10.1007/s11120-005-6966-y