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Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers
Tandem fluorescent protein timers (tFTs) report on protein age through time-dependent change in color, which can be exploited to study protein turnover and trafficking. Each tFT, composed of two fluorescent proteins (FPs) that differ in maturation kinetics, is suited to follow protein dynamics withi...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The American Society for Cell Biology
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4713137/ https://www.ncbi.nlm.nih.gov/pubmed/26609072 http://dx.doi.org/10.1091/mbc.E15-07-0525 |
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author | Khmelinskii, Anton Meurer, Matthias Ho, Chi-Ting Besenbeck, Birgit Füller, Julia Lemberg, Marius K. Bukau, Bernd Mogk, Axel Knop, Michael |
author_facet | Khmelinskii, Anton Meurer, Matthias Ho, Chi-Ting Besenbeck, Birgit Füller, Julia Lemberg, Marius K. Bukau, Bernd Mogk, Axel Knop, Michael |
author_sort | Khmelinskii, Anton |
collection | PubMed |
description | Tandem fluorescent protein timers (tFTs) report on protein age through time-dependent change in color, which can be exploited to study protein turnover and trafficking. Each tFT, composed of two fluorescent proteins (FPs) that differ in maturation kinetics, is suited to follow protein dynamics within a specific time range determined by the maturation rates of both FPs. So far, tFTs have been constructed by combining slower-maturing red fluorescent proteins (redFPs) with the faster-maturing superfolder green fluorescent protein (sfGFP). Toward a comprehensive characterization of tFTs, we compare here tFTs composed of different faster-maturing green fluorescent proteins (greenFPs) while keeping the slower-maturing redFP constant (mCherry). Our results indicate that the greenFP maturation kinetics influences the time range of a tFT. Moreover, we observe that commonly used greenFPs can partially withstand proteasomal degradation due to the stability of the FP fold, which results in accumulation of tFT fragments in the cell. Depending on the order of FPs in the timer, incomplete proteasomal degradation either shifts the time range of the tFT toward slower time scales or precludes its use for measurements of protein turnover. We identify greenFPs that are efficiently degraded by the proteasome and provide simple guidelines for the design of new tFTs. |
format | Online Article Text |
id | pubmed-4713137 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | The American Society for Cell Biology |
record_format | MEDLINE/PubMed |
spelling | pubmed-47131372016-03-30 Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers Khmelinskii, Anton Meurer, Matthias Ho, Chi-Ting Besenbeck, Birgit Füller, Julia Lemberg, Marius K. Bukau, Bernd Mogk, Axel Knop, Michael Mol Biol Cell Articles Tandem fluorescent protein timers (tFTs) report on protein age through time-dependent change in color, which can be exploited to study protein turnover and trafficking. Each tFT, composed of two fluorescent proteins (FPs) that differ in maturation kinetics, is suited to follow protein dynamics within a specific time range determined by the maturation rates of both FPs. So far, tFTs have been constructed by combining slower-maturing red fluorescent proteins (redFPs) with the faster-maturing superfolder green fluorescent protein (sfGFP). Toward a comprehensive characterization of tFTs, we compare here tFTs composed of different faster-maturing green fluorescent proteins (greenFPs) while keeping the slower-maturing redFP constant (mCherry). Our results indicate that the greenFP maturation kinetics influences the time range of a tFT. Moreover, we observe that commonly used greenFPs can partially withstand proteasomal degradation due to the stability of the FP fold, which results in accumulation of tFT fragments in the cell. Depending on the order of FPs in the timer, incomplete proteasomal degradation either shifts the time range of the tFT toward slower time scales or precludes its use for measurements of protein turnover. We identify greenFPs that are efficiently degraded by the proteasome and provide simple guidelines for the design of new tFTs. The American Society for Cell Biology 2016-01-15 /pmc/articles/PMC4713137/ /pubmed/26609072 http://dx.doi.org/10.1091/mbc.E15-07-0525 Text en © 2016 Khmelinskii et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology. |
spellingShingle | Articles Khmelinskii, Anton Meurer, Matthias Ho, Chi-Ting Besenbeck, Birgit Füller, Julia Lemberg, Marius K. Bukau, Bernd Mogk, Axel Knop, Michael Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers |
title | Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers |
title_full | Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers |
title_fullStr | Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers |
title_full_unstemmed | Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers |
title_short | Incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers |
title_sort | incomplete proteasomal degradation of green fluorescent proteins in the context of tandem fluorescent protein timers |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4713137/ https://www.ncbi.nlm.nih.gov/pubmed/26609072 http://dx.doi.org/10.1091/mbc.E15-07-0525 |
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