Spectroscopic method for estimation of MMP-9 enzyme concentration and activity

A. Synak, I. E. Serdiuk, B. Grobelna, R. Fudala, I. Gryczynski, P. Bojarski

Research output: Contribution to journalArticlepeer-review

Abstract

Two highly sensitive fluorescent dye-labelled peptide indicators for metalloproteinase 9 (MMP-9)activity were designed and tested in vitro. Excellent sensitivity of these indicators is achieved by careful selection of the amino acid sequence corresponding to the substrate recognition preference of MMP-9. The phenomenon of Förster resonance energy transfer (FRET)is applied for detection of the peptide cleavage by MMP-9. FRET occurs between fluorescent dyes AMCA (donor)and TAMRA (acceptor)bound to the flexible peptide fragments differing in the donor acceptor separation distance. Enzymatic hydrolysis of the peptide leads to a significant weakening of energy transfer and increased donor fluorescence intensity. The enzyme action efficiency is higher for the longer proline-rich peptide which is reflected in the results of steady-state fluorescence studies. In particular, the use of longer peptide allows shorter detection time of MMP-9 (about 20–30 min). The possible reasons for this effect are discussed.

Original languageEnglish
Article number110936
JournalJournal of Molecular Liquids
Volume286
DOIs
StatePublished - 15 Jul 2019

Keywords

  • Dye-labelled peptide
  • FRET
  • Fluorescence
  • Metalloproteinase 9

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