PMID:1325457
| Citation |
Kessler, D, Herth, W and Knappe, J (1992) Ultrastructure and pyruvate formate-lyase radical quenching property of the multienzymic AdhE protein of Escherichia coli. J. Biol. Chem. 267:18073-9 |
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| Abstract |
The AdhE protein of Escherichia coli is a homopolymer of 96-kDa subunits harboring three Fe(2+)-dependent catalytic functions: acetaldehyde-CoA dehydrogenase, alcohol dehydrogenase, and pyruvate formatelyase (PFL) deactivase. By negative staining electron microscopy, we determined a helical assembly of 20-60 subunits into rods of 45-120 nm in length. The subunit packing is widened along the helix axis when Fe2+ and NAD are present. Chymotrypsin dissects the AdhE polypeptide between Phe762 and Ser763, thereby retaining the alcohol dehydrogenase activity on the NH2-terminal core, but destroying all other activities. PFL deactivation, i.e. quenching of the glycyl radical in PFL by the AdhE protein, was examined with respect to cofactor involvements (Fe2+, NAD, and CoA). This process is coupled to NAD reduction and requires the intact CoA sulfhydryl group. Pyruvate and NADH are inhibitors that affect the steady-state level of the radical form of PFL in a reconstituted interconversion cycle. Studies of cell cultures found that PFL deactivation in situ is initiated at redox potentials of greater than or equal to +100 mV. Our results provide insights into the structure/function organization of the AdhE multienzyme and give a rationale for how its PFL radical quenching activity may be suppressed in situ to enable effective glucose fermentation. |
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| Keywords |
Acetyltransferases/metabolism; Alcohol Dehydrogenase/genetics; Alcohol Dehydrogenase/metabolism; Alcohol Dehydrogenase/ultrastructure; Aldehyde Oxidoreductases/genetics; Aldehyde Oxidoreductases/metabolism; Aldehyde Oxidoreductases/ultrastructure; Chymotrypsin/metabolism; Electron Spin Resonance Spectroscopy; Escherichia coli/enzymology; Escherichia coli Proteins; Free Radicals; Genes, Bacterial; Kinetics; Macromolecular Substances; Microscopy, Electron; Models, Molecular; Molecular Weight; Multienzyme Complexes/genetics; Multienzyme Complexes/metabolism; Multienzyme Complexes/ultrastructure; Peptide Fragments/isolation & purification; Protein Conformation |
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