title: Substrate binding in human indoleamine 2,3-dioxygenase 1: A spectroscopic analysis.
abstract: Human indoleamine 2,3-dioxygenase (hIDO1) is a heme enzyme that catalyzes the oxidative cleavage of the L-tryptophan indole ring. As increased levels of hIDO1 expression in tumor cells correlate with a poor prognosis for surviving several cancer types, hIDO1 has become an appealing drug target for cancer therapy. However, detailed structural knowledge of the catalytically active complex is necessary to eb able to design de novo inhibitors selective for hIDO1. Here we have applied Fourier transform infrared (FTIR) and nanosecond time-resolved optical spectroscopy to hIDO1 variants with modified heme pocket structures to identify important amino acid residues that stabilize the substrate in the active site. A cluster of small side chain residues at positions 260-265 ensures structural flexibility of the binding site. Thr379 and Arg231 are key residues acting in concert to bind the substrate. Thr379 is the final residue of a disordered loop; the neighboring Gly380, however, is still visible in the X-ray structure of the substrate-free protein, 20Å away from the heme iron. Therefore, large-scale conformational changes are necessary to bring Thr379 close to the substrate. The use of substrate analogs further reveals that an indole-like side chain with two aromatic rings and L-stereoisomery at the Cα are required for high affinity binding.
authors.affiliation: Institute of Applied Physics, Karlsruhe Institute of Technology (KIT), Wolfgang-Gaede-Str. 1, 76131, Karlsruhe, Germany.
authors.forename: Karin
authors.initials: K
authors.lastname: Nienhaus
authors.affiliation: Institute of Nanotechnology, and Institute of Toxicology and Genetics, Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany.
authors.forename: Elena
authors.initials: E
authors.lastname: Nickel
authors.affiliation: Institute of Applied Physics, Karlsruhe Institute of Technology (KIT), Wolfgang-Gaede-Str. 1, 76131, Karlsruhe, Germany; Institute of Nanotechnology, and Institute of Toxicology and Genetics, Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany; Department of Physics, University of Illinois at Urbana-Champaign, 1110 West Green Street, Urbana, IL 61801, USA. Electronic address: uli@illinois.edu.
authors.forename: G Ulrich
authors.initials: GU
authors.lastname: Nienhaus
mesh_terms: D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006801:Humans; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase / Q000737:chemistry* / Q000235:genetics; D017433:Protein Structure, Secondary; D017550:Spectroscopy, Fourier Transform Infrared; D013379:Substrate Specificity
chemical_list: C576710:IDO1 protein, human; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase
keywords: Flash photolysis; Fourier transform infrared spectroscopy; Indoleamine 2,3-dioxygenase; Substrate binding