title: Heme Inhibition of [delta]-Aminolevulinic Acid Synthesis Is Enhanced by Glutathione in Cell-Free Extracts of Chlorella.
abstract: In plants, algae, and many bacteria, the heme and chlorophyll precursor, [delta]-aminolevulinic acid (ALA), is synthesized from glutamate in a reaction involving a glutamyl-tRNA intermediate and requiring ATP and NADPH as cofactors. In particulate-free extracts of algae and chloroplasts, ALA synthesis is inhibited by heme. Inclusion of 1.0 mM glutathione (GSH) in an enzyme and tRNA extract, derived from the green alga Chlorella vulgaris, lowered the concentration of heme required for 50% inhibition approximately 10-fold. The effect of GSH could not be duplicated with other reduced sulfhydryl compounds, including mercaptoethanol, dithiothreitol, and cysteine, or with imidazole or bovine serum albumin, which bind to heme and dissociate heme dimers. Absorption spectroscopy indicated that heme was fully reduced in incubation medium containing dithiothreitol, and addition of GSH did not alter the heme reduction state. Oxidized GSH was as effective in enhancing heme inhibition as the reduced form. Co-protoporphyrin IX inhibited ALA synthesis nearly as effectively as heme, and 1.0 mM GSH lowered the concentration required for 50% inhibition approximately 10-fold. Because GSH did not influence the reduction state of heme in the incubation medium, and because GSH could not be replaced by other reduced sulfhydryl compounds or ascorbate, the effect of GSH cannot be explained by action as a sulfhydryl protectant or heme reductant. Preincubation of enzyme extract with GSH, followed by rapid gel filtration, could not substitute for inclusion of GSH with heme during the reaction. The results suggest that GSH must specifically interact with the enzyme extract in the presence of the inhibitor to enhance the inhibition.
authors.affiliation: Department of Biological Sciences, Clemson University, Clemson, South Carolina 29634-1903 (J.D.W., R.W.H., R.D.L., S.Y.G., P.S.B.).
authors.forename: J. D.
authors.initials: JD
authors.lastname: Weinstein
authors.forename: R. W.
authors.initials: RW
authors.lastname: Howell
authors.forename: R. D.
authors.initials: RD
authors.lastname: Leverette
authors.forename: S. Y.
authors.initials: SY
authors.lastname: Grooms
authors.forename: P. S.
authors.initials: PS
authors.lastname: Brignola
authors.forename: S. M.
authors.initials: SM
authors.lastname: Mayer
authors.forename: S. I.
authors.initials: SI
authors.lastname: Beale