Faculté des sciences

Determination of Intracellular Prolyl/Glycyl Proteases in Intact Living Human Cells and Protoporphyrin IX Production as a Reporter System

Berger, Yann ; Chapuis Bernasconi, Catherine ; Schmitt, Frédéric ; Neier, Reinhard ; Juillerat-Jeanneret, Lucienne

In: Chemistry & Biology, 2005, vol. 12, no. 8, p. 867-872

The determination of enzyme activity or inhibition in intact living cells is a problem in the development of inhibitors for intracellular proteases. The production of fluorescent protoporphyrin IX (PpIX) from the nonfluorescent (N)-Gly/Pro-5-aminolevulinic acid (ALA) substrates was used to evaluate the prolyl/glycyl-specific dipeptidylpeptidase IV (DPPIV)-like and prolyloligopeptidase (POP)-like... Plus

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    Summary
    The determination of enzyme activity or inhibition in intact living cells is a problem in the development of inhibitors for intracellular proteases. The production of fluorescent protoporphyrin IX (PpIX) from the nonfluorescent (N)-Gly/Pro-5-aminolevulinic acid (ALA) substrates was used to evaluate the prolyl/glycyl-specific dipeptidylpeptidase IV (DPPIV)-like and prolyloligopeptidase (POP)-like activities of human cells. The results demonstrated that whereas POP-like activity could be attributed to the actual POP, the DPPIV-like activity could be related to actual DPPIV only in one colon cell line. In the other breast and colon cell lines, DPPIV-like activity was intracellular and displayed by other prolyl-specific aminopeptidases. Our experiments also demonstrated the involvement of gylcyl-specific proteases in the processing of ALA precursors. These observations have important consequences for the development and evaluation of selective inhibitors for these enzymes.