Print Email Facebook Twitter A traffic light enzyme Title A traffic light enzyme: acetate binding reversibly switches chlorite dismutase from a red- to a green-colored heme protein Author Mahor, D. (TU Delft BT/Biocatalysis) Puschmann, J. (TU Delft BT/Biocatalysis) van den Haak, Menno (Student TU Delft) Kooij, Pepijn J. (Student TU Delft) van den Ouden, David L.J. (Student TU Delft) Strampraad, M.J.F. (TU Delft BT/Biocatalysis) Srour, B. (TU Delft BT/Biocatalysis) Hagedoorn, P.L. (TU Delft BT/Biocatalysis) Date 2020 Abstract Abstract: Chlorite dismutase is a unique heme enzyme that catalyzes the conversion of chlorite to chloride and molecular oxygen. The enzyme is highly specific for chlorite but has been known to bind several anionic and neutral ligands to the heme iron. In a pH study, the enzyme changed color from red to green in acetate buffer pH 5.0. The cause of this color change was uncovered using UV–visible and EPR spectroscopy. Chlorite dismutase in the presence of acetate showed a change of the UV–visible spectrum: a redshift and hyperchromicity of the Soret band from 391 to 404 nm and a blueshift of the charge transfer band CT1 from 647 to 626 nm. Equilibrium binding titrations with acetate resulted in a dissociation constant of circa 20 mM at pH 5.0 and 5.8. EPR spectroscopy showed that the acetate bound form of the enzyme remained high spin S = 5/2, however with an apparent change of the rhombicity and line broadening of the spectrum. Mutagenesis of the proximal arginine Arg183 to alanine resulted in the loss of the ability to bind acetate. Acetate was discovered as a novel ligand to chlorite dismutase, with evidence of direct binding to the heme iron. The green color is caused by a blueshift of the CT1 band that is characteristic of the high spin ferric state of the enzyme. Any weak field ligand that binds directly to the heme center may show the red to green color change, as was indeed the case for fluoride. Subject Acetate bindingCharge transfer bandChlorite dismutaseElectron paramagnetic resonance (EPR)Green heme protein To reference this document use: http://resolver.tudelft.nl/uuid:afeb61ec-58fc-4d5c-8fb8-a016040ae543 DOI https://doi.org/10.1007/s00775-020-01784-1 ISSN 0949-8257 Source Journal of Biological Inorganic Chemistry, 25 (4), 609-620 Part of collection Institutional Repository Document type journal article Rights © 2020 D. Mahor, J. Puschmann, Menno van den Haak, Pepijn J. Kooij, David L.J. van den Ouden, M.J.F. Strampraad, B. Srour, P.L. Hagedoorn Files PDF Mahor2020_Article_ATraffi ... teBind.pdf 2.79 MB Close viewer /islandora/object/uuid:afeb61ec-58fc-4d5c-8fb8-a016040ae543/datastream/OBJ/view