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Protein Eng. 1999 Dec;12(12):1035-40.

Attenuation of green fluorescent protein half-life in mammalian cells.

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1
CRC Chromosome Molecular Biology Group, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.

Abstract

The half-life of the green fluorescent protein (GFP) was determined biochemically in cultured mouse LA-9 cells. The wild-type protein was found to be stable with a half-life of approximately 26 h, but could be destabilized by the addition of putative proteolytic signal sequences derived from proteins with shorter half-lives. A C-terminal fusion of a PEST sequence from the mouse ornithine decarboxylase gene reduced the half-life to 9.8 h, resulting in a GFP variant suitable for the study of dynamic cellular processes. In an N-terminal fusion containing the mouse cyclin B1 destruction box, it was reduced to 5.8 h, with most degradation taking place at metaphase. The combination of both sequences produced a similar GFP half-life of 5.5 h. Thus, the stability of this marker protein can be controlled in predetermined ways by addition of the appropriate proteolytic signals.

PMID:
10611396
[Indexed for MEDLINE]
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