Synthesis of the Novel Covalent Cysteine Proteases Inhibitor with Iodoacetic Functional Group

Molecules. 2020 Feb 13;25(4):813. doi: 10.3390/molecules25040813.

Abstract

This work presents the synthesis of the novel covalent inhibitor of cysteine proteases where epoxide has been replaced by the iodoacetyl functional group. The molecule, similar in action to E-64 and DCG-04, the commonly applied inhibitors, is additionally biotinylated and contains tyrosyl iodination sites. The Fmoc solid phase synthesis has been applied. Conjugation of iodoacetic acid with the peptide was optimized by testing different conjugation agents. The purity of the final product was verified by mass spectrometry and its bioactivity was tested by incubation with a model cysteine protease-staphopain C. Finally, it was shown that the synthesized inhibitor binds to the protein at the ratio of 1:1. More detailed analysis by means of tandem mass spectrometry proved that the inhibitor binds to the cysteine present in the active site of the enzyme.

Keywords: DCG-04; cysteine protease; inhibitor; iodoacetic acid; solid phase synthesis.

MeSH terms

  • Biotinylation
  • Cysteine Endopeptidases / chemical synthesis*
  • Cysteine Endopeptidases / chemistry*
  • Cysteine Proteinase Inhibitors / chemical synthesis*
  • Cysteine Proteinase Inhibitors / chemistry*
  • Iodoacetic Acid / chemistry*
  • Leucine / analogs & derivatives
  • Leucine / chemistry
  • Molecular Structure
  • Solid-Phase Synthesis Techniques / methods
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Substances

  • Cysteine Proteinase Inhibitors
  • DCG 04
  • Cysteine Endopeptidases
  • Leucine
  • Iodoacetic Acid