Evolutionary Optimization of Fluorescent Proteins for Intracellular FRET  

Oleh:

Nguyen, Annalee W. ; Daugherty, Patrick S.

Jenis: Article from Journal - ilmiah internasional
Dalam koleksi: Nature Biotechnology: The Science and Business of Biotechnology vol. 23 no. 3 (Mar. 2005), page 355-360.
Topik: proteins; evolutionary optimization; fluorescent proteins; intracellular FRET

Ketersediaan

Perpustakaan Pusat (Semanggi) Nomor Panggil: NN9.2 Non-tandon: 1 (dapat dipinjam: 0) Tandon: tidak ada

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Isi artikel

Fluorescent proteins that exhibit forster resonance energy transfer (FRET) have mad a strong impact as they enable measurement of molecular scale distances through changes in fluorescence. FRET based approaches have enabled otherwise intractable measurements of molecular concentrations, binding interactions and catalytic activity but are limited by the dynamic range and sensitivity of the donor acceptor pair. To address this problem, we applied a quantitative evolutionary strategy using fluorescence activated cell sorting to optimize a cyan yellow fluorescent protein pair for FRET. the resulting pair, CyPet-YPet, exhibited a 20-fold ratiometric FRET signal change, as compared to threefold for the parental pair. The optimized FRET pair enable high throughput flow cytometric screening of cells undergoing caspase 3-dependent apoptosis. The CyPet-YPet energy transger pair provides substantially improved sensitivity and dynamic range for a broad range of molecular imaging and screening applications.

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