Biochemical and crystallographic investigations into isonitrile formation by a nonheme iron-dependent oxidase/decarboxylase
| Autor: | Maanasa Narayanamoorthy, Chaoxiang Ren, Antonio Del Rio Flores, Jan Paulo T. Zaragoza, Wenjun Zhang, Heather J. Kulik, Wenlong Cai, Rimsha Mehmood, Rohan Jonnalagadda, Catherine L. Drennan |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Models Molecular Protein Conformation alpha-Helical Protein Conformation Gene Expression Crystallography X-Ray Biochemistry Medical and Health Sciences Substrate Specificity Dioxygenase Models structure function Moiety Cloning Molecular INLP isonitrile lipopeptide biology Chemistry Aminobutyrates Stereoisomerism mononuclear Fe(II) α-ketoglutarate dependent dioxygenase Biological Sciences Condensation reaction Enzyme structure Recombinant Proteins Streptomyces stoichiometry LC-HRMS liquid chromatography–high-resolution mass spectrometry 5.1 Pharmaceuticals Ketoglutaric Acids INBA (R)-3-isocyano butanoic acid Development of treatments and therapeutic interventions RESP restrained electrostatic potential Protein Binding Research Article Biochemistry & Molecular Biology Stereochemistry Iron Genetic Vectors Glycine Arginine Cofactor Adduct Dioxygenases 03 medical and health sciences Bacterial Proteins CABA (R)-3-((carboxylmethyl)amino)butanoic acid Nitriles Escherichia coli Histidine Protein Interaction Domains and Motifs enzyme mechanism Binding site crystallography Molecular Biology Binding Sites 030102 biochemistry & molecular biology alpha-Helical Active site Molecular Cell Biology molecular dynamics 030104 developmental biology Chemical Sciences biology.protein X-Ray beta-Strand Protein Conformation beta-Strand Vanadates Cloning |
| Zdroj: | The Journal of Biological Chemistry Elsevier |
| ISSN: | 1083-351X |
| Popis: | 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 (1.45-1.96Å 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 10Å 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. |
| Databáze: | OpenAIRE |
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