title: Asp274 and his346 are essential for heme binding and catalytic function of human indoleamine 2,3-dioxygenase.
abstract: L-Tryptophan is the least abundant essential amino acid in humans. Indoleamine 2,3-dioxgyenase (IDO) is a cytosolic heme protein which, together with the hepatic enzyme tryptophan 2,3-dioxygenase, catalyzes the first and rate-limiting step in the major pathway of tryptophan metabolism, the kynurenine pathway. The physiological role of IDO is not fully understood but is of great interest, because IDO is widely distributed in human tissues, can be up-regulated via cytokines such as interferon-gamma, and can thereby modulate the levels of tryptophan, which is vital for cell growth. To identify which amino acid residues are important in substrate or heme binding in IDO, site-directed mutagenesis of conserved residues in the IDO gene was undertaken. Because it had been proposed that a histidine residue might be the proximal heme ligand in IDO, mutation to alanine of the three highly conserved histidines His16, His303, and His346 was conducted. Of these, only His346 was shown to be essential for heme binding, indicating that this histidine residue may be the proximal ligand and suggesting that neither His303 nor His16 act as the proximal ligand. Site-directed mutagenesis of Asp274 also compromised the ability of IDO to bind heme. This observation indicates that Asp274 may coordinate to heme directly as the distal ligand or is essential in maintaining the conformation of the heme pocket.
authors.affiliation: Australian Cataract Research Foundation, Department of Chemistry, University of Wollongong, New South Wales, Australia.
authors.forename: Tamantha K
authors.initials: TK
authors.lastname: Littlejohn
authors.forename: Osamu
authors.initials: O
authors.lastname: Takikawa
authors.forename: Roger J W
authors.initials: RJ
authors.lastname: Truscott
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Walker
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000737:chemistry*; D015153:Blotting, Western; D002384:Catalysis; D002455:Cell Division; D002942:Circular Dichroism; D016207:Cytokines / Q000096:biosynthesis; D004578:Electron Spin Resonance Spectroscopy; D004591:Electrophoresis, Polyacrylamide Gel; D006418:Heme / Q000737:chemistry*; D006639:Histidine / Q000737:chemistry*; D006801:Humans; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D007371:Interferon-gamma / Q000096:biosynthesis; D007700:Kinetics; D008024:Ligands; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D017386:Sequence Homology, Amino Acid; D014364:Tryptophan / Q000737:chemistry; D014366:Tryptophan Oxygenase / Q000737:chemistry*; D014466:Ultraviolet Rays; D015854:Up-Regulation
chemical_list: D016207:Cytokines; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D008024:Ligands; D001224:Aspartic Acid; D006418:Heme; D006639:Histidine; D007371:Interferon-gamma; D014364:Tryptophan; D014366:Tryptophan Oxygenase; D000409:Alanine