Probing the cathepsin D using a BODIPY FL–pepstatin A: applications in fluorescence polarization and microscopy
作者： Chii-Shiarng Chen , Wan-Nan U Chen , Mingjie Zhou , Seksiri Arttamangkul , Richard P Haugland
DOI: 10.1016/S0165-022X(00)00048-8
关键词:
摘要: Redistribution of cathepsin D, a major lysosomal aspartic endopeptidase, has been related to various pathological progressions during tumor formation and oxidation stress. We have synthesized fluorescent probe for where the pepstatin A was covalently conjugated with BODIPY (Boron dipyrromethene difluoride) fluorophore. In vitro, FL-pepstatin inhibits D activity an IC50 10 nM. The nature its binding further characterized using fluorescence polarization measurement. Results showed that selectively binds at pH 4.5. fixed cells, stained lysosomes, it co-localized D. This staining depleted when cells were co-incubated unlabeled in acidic buffer. live is internalized transported lysosomes. lysosomes can be competed A. These properties, along good photostability FL fluorophore, make this novel tool study secretion trafficking
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