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PMID:8325851

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Citation

Flint, DH, Emptage, MH, Finnegan, MG, Fu, W and Johnson, MK (1993) The role and properties of the iron-sulfur cluster in Escherichia coli dihydroxy-acid dehydratase. J. Biol. Chem. 268:14732-42

Abstract

Dihydroxy-acid dehydratase has been purified from Escherichia coli and characterized as a homodimer with a subunit molecular weight of 66,000. The combination of UV visible absorption, EPR, magnetic circular dichroism, and resonance Raman spectroscopies indicates that the native enzyme contains a [4Fe-4S]2+,+ cluster, in contrast to spinach dihydroxy-acid dehydratase which contains a [2Fe-2S]2+,+ cluster (Flint, D. H., and Emptage, M. H. (1988) J. Biol. Chem. 263, 3558-3564). In frozen solution, the reduced [4Fe-4S]+ cluster has a S = 3/2 ground state with minor contributions from forms with S = 1/2 and possibly S = 5/2 ground states. Resonance Raman studies of the [4Fe-4S]2+ cluster in E. coli dihydroxy-acid dehydratase indicate non-cysteinyl coordination of a specific iron, which suggests that it is likely to be directly involved in catalysis as is the case with aconitase (Emptage, M. H., Kent, T. A., Kennedy, M. C., Beinert, H., and Münck, E. (1983) Proc. Natl. Acad. Sci. U.S.A. 80, 4674-4678). Dihydroxy-acid dehydratase from E. coli is inactivated by O2 in vitro and in vivo as a result of oxidative degradation of the [4Fe-4S]cluster. Compared to aconitase, the oxidized cluster of E. coli dihydroxy-acid dehydratase appears to be less stable as either a cubic or linear [3Fe-4S] cluster or a [2Fe-2S] cluster. Oxidative degradation appears to lead to a complete breakdown of the Fe-S cluster, and the resulting protein cannot be reactivated with Fe2+ and thiol reducing agents.

Links

PubMed

Keywords

Amino Acid Sequence; Enzyme Activation; Escherichia coli/enzymology; Hydro-Lyases/antagonists & inhibitors; Hydro-Lyases/chemistry; Hydro-Lyases/isolation & purification; Hydro-Lyases/metabolism; Iron-Sulfur Proteins/analysis; Iron-Sulfur Proteins/metabolism; Molecular Sequence Data; Oxidation-Reduction; Spectrum Analysis

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

ECOLI:ILVD

acts_upstream_of_or_within

GO:0009099: valine biosynthetic process

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete

ECOLI:ILVD

involved_in

GO:0009099: valine biosynthetic process

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete

ECOLI:ILVD

enables

GO:0051539: 4 iron, 4 sulfur cluster binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

ECOLI:ILVD

enables

GO:0051536: iron-sulfur cluster binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

ECOLI:ILVD

enables

GO:0004160: dihydroxy-acid dehydratase activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

See also

References

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