Irregularities in enzyme assays: The case of macrophage migration inhibitory factor

Bioorg Med Chem Lett. 2016 Jun 15;26(12):2764-2767. doi: 10.1016/j.bmcl.2016.04.074. Epub 2016 Apr 26.

Abstract

Inhibitors of human macrophage migration inhibitory factor (MIF) previously reported in the literature have been reexamined by synthesis, assaying for tautomerase activity, and protein crystallography. Substantial inconsistencies between prior and current assay results are noted. They appear to arise from difficulties with the tautomerase substrates, solubility issues, and especially covalent inhibition. Incubation time variation shows that 3, 4, 6, and 9 are covalent or slow-binding inhibitors. Two protein crystal structures are provided; one confirms that the twice-discovered 3 is a covalent inhibitor.

Keywords: MIF; Protein crystallography; Reproducibility; Tautomerase assay.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Dose-Response Relationship, Drug
  • Enzyme Inhibitors / chemical synthesis
  • Enzyme Inhibitors / chemistry
  • Enzyme Inhibitors / pharmacology*
  • Humans
  • Intramolecular Oxidoreductases / antagonists & inhibitors*
  • Intramolecular Oxidoreductases / metabolism
  • Macrophage Migration-Inhibitory Factors / antagonists & inhibitors*
  • Macrophage Migration-Inhibitory Factors / metabolism
  • Models, Molecular
  • Molecular Structure
  • Structure-Activity Relationship

Substances

  • Enzyme Inhibitors
  • Macrophage Migration-Inhibitory Factors
  • Intramolecular Oxidoreductases
  • MIF protein, human