Publications

• Evidence for catalytic intermediates involved in generating the chromopyrrolic acid scaffold of rebeccamycin by RebO and RebD.

  Spolitak T., Ballou D.P.

  We provide the first experimental evidence for intermediates being involved in catalysis by RebD in generating the chromopyrrolic acid (CPA) scaffold of rebeccamycin. In the presence of its substrates (indole pyruvate imine - IPAI - and H2O2 both produced by the flavoprotein oxidase RebO that oxid ... >> More

  We provide the first experimental evidence for intermediates being involved in catalysis by RebD in generating the chromopyrrolic acid (CPA) scaffold of rebeccamycin. In the presence of its substrates (indole pyruvate imine - IPAI - and H2O2 both produced by the flavoprotein oxidase RebO that oxidizes tryptophan), RebD reacts as a peroxidase forming two IPAI radicals that recombine as a C-C bond in the CPA. When catalase is included to remove H2O2, CPA can still be formed because the IPAI rapidly reduces RebD, which reacts with O2, utilizing oxidase-peroxidase chemistry to produce CPA. Reduced RebD can also react with H2O2 forming Cpd II directly, which can oxidize IPAI. Stopped-flow spectrophotometric studies demonstrated that during the reaction of RebO and RebD with Trp and oxygen, a species with a red-shifted Soret band at 424.5 nm appeared. This species can react with either guaiacol or ABTS to form ferric RebD, suggesting that it is Cpd II of RebD involved in the formation of CPA. In summary, the studies reveal new and unusual aspects peroxidase and peroxygenase chemistry used by RebD in catalyzing carbon-carbon oxidative coupling reactions that are involved in biosynthesis of indolocarbazoles. << Less

  Arch. Biochem. Biophys. 573:111-119(2015) [PubMed] [EuropePMC]

  This publication is cited by 6 other entries.

• Enzymatic generation of the chromopyrrolic acid scaffold of rebeccamycin by the tandem action of RebO and RebD.

  Howard-Jones A.R., Walsh C.T.

  During the biosynthesis of the fused six-ring indolocarbazole scaffolds of rebeccamycin and staurosporine, two molecules of L-tryptophan are processed to a pyrrole-containing five-ring intermediate known as chromopyrrolic acid. We report here the heterologous expression of RebO and RebD from the r ... >> More

  During the biosynthesis of the fused six-ring indolocarbazole scaffolds of rebeccamycin and staurosporine, two molecules of L-tryptophan are processed to a pyrrole-containing five-ring intermediate known as chromopyrrolic acid. We report here the heterologous expression of RebO and RebD from the rebeccamycin biosynthetic pathway in Escherichia coli, and tandem action of these two enzymes to construct the dicarboxypyrrole ring of chromopyrrolic acid. Chromopyrrolic acid is oxidized by six electrons compared to the starting pair of L-tryptophan molecules. RebO is an L-tryptophan oxidase flavoprotein and RebD a heme protein dimer with both catalase and chromopyrrolic acid synthase activity. Both enzymes require dioxygen as a cosubstrate. RebD on its own is incompetent with L-tryptophan but will convert the imine of indole-3-pyruvate to chromopyrrolic acid. It displays a substrate preference for two molecules of indole-3-pyruvic acid imine, necessitating a net two-electron oxidation to give chromopyrrolic acid. << Less

  Biochemistry 44:15652-15663(2005) [PubMed] [EuropePMC]

  This publication is cited by 8 other entries.

• Enzymatic assembly of the bis-indole core of rebeccamycin.

  Nishizawa T., Gruschow S., Jayamaha D.H., Nishizawa-Harada C., Sherman D.H.

  Rebeccamycin is a member of the family of indolocarbazole antibiotics with broad spectrum antitumor activity. The indolocarbazole framework is derived from two molecules of tryptophan, but very little is known about the enzymes involved in rebeccamycin biosynthesis. Here, we show that RebD is resp ... >> More

  Rebeccamycin is a member of the family of indolocarbazole antibiotics with broad spectrum antitumor activity. The indolocarbazole framework is derived from two molecules of tryptophan, but very little is known about the enzymes involved in rebeccamycin biosynthesis. Here, we show that RebD is responsible for all catalytic steps forming the central pyrrole ring of chlorochromopyrrolic acid from two molecules of chloroindolepyruvic acid. This transformation does not require any additional cofactors and constitutes the first step of bis-indole formation in the biosynthesis of rebeccamycin. << Less

  J Am Chem Soc 128:724-725(2006) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.