title: Targeting distinct tautomerase sites of D-DT and MIF with a single molecule for inhibition of neutrophil lung recruitment.
abstract: We report a new inflammatory activity for extracellular d-dopachrome tautomerase (D-DT), the recruitment of neutrophils to the lung on D-DT intratracheal installation of C57BL/6J mice with an EC50 of 5.6 μg. We also find that D-DT and macrophage migration inhibitory factor (MIF) have additive effects in neutrophil recruitment. Although the tautomerase site of D-DT and its homologue MIF are biophysically very different, 4-iodo-6-phenylpyrimidine (4-IPP) forms a covalent bond with Pro-1 of both proteins, resulting in a 6-phenylpyrimidine (6-PP) adduct. Recruitment of neutrophils to the lung for the 6-PP adducts of D-DT and MIF are reduced by ∼ 50% relative to the apo proteins, demonstrating that an unmodified Pro-1 is important for this activity, but there is no cooperativity in inhibition of the proteins together. The differences in the binding mode of the 6-PP adduct for D-DT was determined by crystallographic studies at 1.13 Å resolution and compared to the structure of the MIF-6-PP complex. There are major differences in the location of the 6-PP adduct to the D-DT and MIF active sites that provide insight into the lack of cooperativity by 4-IPP and into tuning the properties of the covalent inhibitors of D-DT and MIF that are necessary for the development of therapeutic small molecules against neutrophil damage from lung infections such as Pseudomonas aeruginosa in cystic fibrosis and immunocompromised patients.
authors.affiliation: Department of Pharmacology.
authors.forename: Deepa
authors.initials: D
authors.lastname: Rajasekaran
authors.affiliation: Department of Pharmacology.
authors.forename: Swen
authors.initials: S
authors.lastname: Zierow
authors.affiliation: Department of Pediatrics.
authors.forename: Mansoor
authors.initials: M
authors.lastname: Syed
authors.affiliation: Department of Internal Medicine, and Yale Cancer Center, Yale University, New Haven, Connecticut, USA.
authors.forename: Richard
authors.initials: R
authors.lastname: Bucala
authors.affiliation: Department of Pediatrics.
authors.forename: Vineet
authors.initials: V
authors.lastname: Bhandari
authors.affiliation: Department of Pharmacology, Yale Cancer Center, Yale University, New Haven, Connecticut, USA elias.lolis@yale.edu.
authors.forename: Elias J
authors.initials: EJ
authors.lastname: Lolis
mesh_terms: D000818:Animals; D020134:Catalytic Domain / Q000187:drug effects*; D002465:Cell Movement; D019746:Intramolecular Oxidoreductases / Q000378:metabolism*; D008168:Lung / Q000187:drug effects / Q000378:metabolism*; D008263:Macrophage Migration-Inhibitory Factors / Q000378:metabolism*; D008810:Mice, Inbred C57BL; D008958:Models, Molecular; D020556:Neutrophil Infiltration / Q000187:drug effects*; D009504:Neutrophils / Q000166:cytology*; D011743:Pyrimidines / Q000494:pharmacology*
chemical_list: C532251:4-iodo-6-phenylpyrimidine; D008263:Macrophage Migration-Inhibitory Factors; D011743:Pyrimidines; D019746:Intramolecular Oxidoreductases; C076231:Mif protein, mouse; C042501:dopachrome isomerase
keywords: 4-IPP; covalent inhibitor