Biochemical and in vivo characterization of a small, membrane-permeant, caspase-activatable far-red fluorescent peptide for imaging apoptosis

Biochemistry. 2007 Apr 3;46(13):4055-65. doi: 10.1021/bi061959n. Epub 2007 Mar 10.


Apoptosis is an important process involved in diverse developmental pathways, homeostasis, and response to therapy for a variety of diseases. Thus, noninvasive methods to study regulation and to monitor cell death in cells and whole animals are desired. To specifically detect apoptosis in vivo, a novel cell-permeable activatable caspase substrate, TcapQ647, was synthesized and Km, kcat, and Ki values were biochemically characterized. Specific cleavage of TcapQ647 by effector caspases was demonstrated using a panel of purified recombinant enzyme assays. Of note, caspase 3 was shown to cleave TcapQ647 with a kcat 7-fold greater than caspase 7 and 16-fold greater than caspase 6. No evidence of TcapQ647 cleavage by initiator caspases was observed. In KB 3-1 or Jurkat cells treated with cytotoxic agents or C6-ceramide, TcapQ647 detected apoptosis in individual- and population-based fluorescent cell assays in an effector caspase inhibitor-specific manner. Further, only background fluorescence was observed in cells incubated with dTcapQ647, a noncleavable all d-amino acid control peptide. Finally, in vivo experiments demonstrated the utility of TcapQ647 to detect parasite-induced apoptosis in human colon xenograft and liver abscess mouse models. Thus, TcapQ647 represents a sensitive, effector caspase-specific far-red "smart" probe to noninvasively monitor apoptosis in vivo.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Apoptosis / physiology
  • Blotting, Western
  • Caspases / metabolism*
  • Colonic Neoplasms
  • Cyclic AMP / analogs & derivatives
  • Cyclic AMP / chemistry
  • Fluorescent Dyes / chemical synthesis*
  • Fluorescent Dyes / chemistry*
  • HeLa Cells
  • Humans
  • In Situ Nick-End Labeling
  • Jurkat Cells
  • Kinetics
  • Liver / metabolism
  • Mice
  • Neoplasm Transplantation
  • Peptides / chemical synthesis*
  • Peptides / chemistry*
  • Spectrometry, Fluorescence
  • Transplantation, Heterologous


  • Fluorescent Dyes
  • Peptides
  • TcapQ647 peptide
  • 8-aminohexylamino cAMP
  • Cyclic AMP
  • Caspases