Luminescent quantum dots fluorescence resonance energy transfer-based probes for enzymatic activity and enzyme inhibitors.
The paper describes the development and characterization of analytical properties of quantum dot-based probes for enzymatic activity and for screening enzyme inhibitors. The luminescent probes are based on fluorescence resonance energy transfer (FRET) between luminescent quantum dots that serve as donors and rhodamine acceptors that are immobilized to the surface of the quantum dots through peptide linkers. Peptide-coated CdSe/ZnS quantum dots were prepared using a one-step ligand exchange process in which RGDC peptide molecules replace trioctylphosphine oxide (TOPO) molecules as the capping ligands of the quantum dots. The peptide molecules were bound to the surface of the CdSe/ZnS quantum dots through the thiol group of the peptide cysteine residue. The peptide-coated quantum dots were labeled with rhodamine to form the FRET probes. The emission quantum yield of the quantum dot FRET probes was 4-fold lower than the emission quantum yield of TOPO-capped quantum dots. However, the quantum dot FRET probes were sufficiently bright to enable quantitative enzyme and enzyme inhibition assays. The probes were used first to test the enzymatic activity of trypsin in solution based on FRET signal changes of the quantum dot-based enzymatic probes in the presence of proteolytic enzymes. For example, exposure of the quantum dot FRET probes to 500 microg/mL trypsin for 15 min resulted in 60% increase in the photoluminescence of the quantum dots and a corresponding decrease in the emission of the rhodamine molecules. These changes resulted from the release of rhodamine molecules from the surface of the quantum dots due to enzymatic cleavage of the peptide molecules. The quantum dot FRET-based probes were used to monitor the enzymatic activity of trypsin and to screen trypsin inhibitors for their inhibition efficiency.
Shi
Lifang
L
Department of Chemistry and the Advanced Material Research Institute, University of New Orleans, New Orleans, Louisiana 70148, USA.
Rosenzweig
Nitsa
N
Rosenzweig
Zeev
Z
D019187:Cadmium Compounds / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000032:analysis / Q000378:metabolism; D004798:Enzymes / Q000032:analysis / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry; D008163:Luminescent Measurements / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D009943:Organophosphorus Compounds / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D045663:Quantum Dots; D012235:Rhodamines / Q000737:chemistry; D018036:Selenium Compounds / Q000737:chemistry; D012680:Sensitivity and Specificity; D013440:Sulfides / Q000737:chemistry; D014357:Trypsin / Q000032:analysis / Q000378:metabolism; D017967:Zinc Compounds / Q000737:chemistry
D019187:Cadmium Compounds; D004791:Enzyme Inhibitors; D004798:Enzymes; D005456:Fluorescent Dyes; D009943:Organophosphorus Compounds; D010455:Peptides; D012235:Rhodamines; D018036:Selenium Compounds; D013440:Sulfides; D017967:Zinc Compounds; C044965:trioctyl phosphine oxide; C058667:cadmium selenide; D014357:Trypsin; C031238:zinc sulfide