Silica microspheres functionalized with the iminodiacetic acid/copper(II) complex as a peroxidase mimic for use in metal affinity chromatography-based colorimetric determination of histidine-tagged proteins

Gokcal B. , Kip C., Sahinbas D., Celik E. , Tuncel A.

MICROCHIMICA ACTA, vol.187, no.2, 2020 (Journal Indexed in SCI) identifier identifier identifier

  • Publication Type: Article / Article
  • Volume: 187 Issue: 2
  • Publication Date: 2020
  • Doi Number: 10.1007/s00604-019-4087-0
  • Title of Journal : MICROCHIMICA ACTA


Monodisperse porous silica microspheres were functionalized with the iminodiacetic acid/copper(II) complex and then evaluated as a group-specific peroxidase-mimicking nanozyme for colorimetric determination of histidine-tagged (His-tagged) proteins. The green fluorescent protein (GFP) was selected as a typical His-tagged protein. The specificity for GFP and the peroxidase-like activity for the selected substrate were obtained by immobilizing the complex on the porous microspheres. The modified microspheres were also evaluated as a group specific immobilized metal affinity chromatography (IMAC) sorbent for the purification of GFP from Escherichia coli extract. The peroxidase-like activity of the microspheres was inhibited by the GFP adsorbed onto the microspheres due to the interaction of His-tagged protein with the immobilized Cu(II) complex. Ortho-phenylenediamine is used as a substrate for the enzyme mimic. The photometric response (measured at 416 nm) is linear in the 9.0-92 mu g center dot mL(-1) GFP concentration range in E. coli lysate. The limit of detection is 6.9 mu g center dot mL(-1).