Publications

• Isonitrile Formation by a Non-Heme Iron(II)-Dependent Oxidase/Decarboxylase.

  Harris N.C., Born D.A., Cai W., Huang Y., Martin J., Khalaf R., Drennan C.L., Zhang W.

  The electron-rich isonitrile is an important functionality in bioactive natural products, but its biosynthesis has been restricted to the IsnA family of isonitrile synthases. We herein provide the first structural and biochemical evidence of an alternative mechanism for isonitrile formation. ScoE, ... >> More

  The electron-rich isonitrile is an important functionality in bioactive natural products, but its biosynthesis has been restricted to the IsnA family of isonitrile synthases. We herein provide the first structural and biochemical evidence of an alternative mechanism for isonitrile formation. ScoE, a putative non-heme iron(II)-dependent enzyme from Streptomyces coeruleorubidus, was shown to catalyze the conversion of (R)-3-((carboxymethyl)amino)butanoic acid to (R)-3-isocyanobutanoic acid through an oxidative decarboxylation mechanism. This work further provides a revised scheme for the biosynthesis of a unique class of isonitrile lipopeptides, of which several members are critical for the virulence of pathogenic mycobacteria. << Less

  Angew. Chem. Int. Ed. Engl. 57:9707-9710(2018) [PubMed] [EuropePMC]

  This publication is cited by 15 other entries.

• Biosynthesis of isonitrile lipopeptides by conserved nonribosomal peptide synthetase gene clusters in Actinobacteria.

  Harris N.C., Sato M., Herman N.A., Twigg F., Cai W., Liu J., Zhu X., Downey J., Khalaf R., Martin J., Koshino H., Zhang W.

  A putative lipopeptide biosynthetic gene cluster is conserved in many species of Actinobacteria, including <i>Mycobacterium tuberculosis</i> and <i>M. marinum</i>, but the specific function of the encoding proteins has been elusive. Using both in vivo heterologous reconstitution and in vitro bioch ... >> More

  A putative lipopeptide biosynthetic gene cluster is conserved in many species of Actinobacteria, including <i>Mycobacterium tuberculosis</i> and <i>M. marinum</i>, but the specific function of the encoding proteins has been elusive. Using both in vivo heterologous reconstitution and in vitro biochemical analyses, we have revealed that the five encoding biosynthetic enzymes are capable of synthesizing a family of isonitrile lipopeptides (INLPs) through a thio-template mechanism. The biosynthesis features the generation of isonitrile from a single precursor Gly promoted by a thioesterase and a nonheme iron(II)-dependent oxidase homolog and the acylation of both amino groups of Lys by the same isonitrile acyl chain facilitated by a single condensation domain of a nonribosomal peptide synthetase. In addition, the deletion of INLP biosynthetic genes in <i>M. marinum</i> has decreased the intracellular metal concentration, suggesting the role of this biosynthetic gene cluster in metal transport. << Less

  Proc. Natl. Acad. Sci. U.S.A. 114:7025-7030(2017) [PubMed] [EuropePMC]

  This publication is cited by 20 other entries.

• Biochemical and crystallographic investigations into isonitrile formation by a nonheme iron-dependent oxidase/decarboxylase.

  Jonnalagadda R., Del Rio Flores A., Cai W., Mehmood R., Narayanamoorthy M., Ren C., Zaragoza J.P.T., Kulik H.J., Zhang W., Drennan C.L.

  The isonitrile moiety is found in marine sponges and some microbes, where it plays a role in processes such as virulence and metal acquisition. Until recently only one route was known for isonitrile biosynthesis, a condensation reaction that brings together a nitrogen atom of l-Trp/l-Tyr with a ca ... >> More

  The isonitrile moiety is found in marine sponges and some microbes, where it plays a role in processes such as virulence and metal acquisition. Until recently only one route was known for isonitrile biosynthesis, a condensation reaction that brings together a nitrogen atom of l-Trp/l-Tyr with a carbon atom from ribulose-5-phosphate. With the discovery of ScoE, a mononuclear Fe(II) α-ketoglutarate-dependent dioxygenase from Streptomyces coeruleorubidus, a second route was identified. ScoE forms isonitrile from a glycine adduct, with both the nitrogen and carbon atoms coming from the same glycyl moiety. This reaction is part of the nonribosomal biosynthetic pathway of isonitrile lipopeptides. Here, we present structural, biochemical, and computational investigations of the mechanism of isonitrile formation by ScoE, an unprecedented reaction in the mononuclear Fe(II) α-ketoglutarate-dependent dioxygenase superfamily. The stoichiometry of this enzymatic reaction is measured, and multiple high-resolution (1.45-1.96 Å resolution) crystal structures of Fe(II)-bound ScoE are presented, providing insight into the binding of substrate, (R)-3-((carboxylmethyl)amino)butanoic acid (CABA), cosubstrate α-ketoglutarate, and an Fe(IV)=O mimic oxovanadium. Comparison to a previously published crystal structure of ScoE suggests that ScoE has an "inducible" α-ketoglutarate binding site, in which two residues arginine-157 and histidine-299 move by approximately 10 Å from the surface of the protein into the active site to create a transient α-ketoglutarate binding pocket. Together, data from structural analyses, site-directed mutagenesis, and computation provide insight into the mode of α-ketoglutarate binding, the mechanism of isonitrile formation, and how the structure of ScoE has been adapted to perform this unusual chemical reaction. << Less

  J. Biol. Chem. 296:100231-100231(2021) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.