Journal
JOURNAL OF PROTEOME RESEARCH
Volume 11, Issue 1, Pages 476-486Publisher
AMER CHEMICAL SOC
DOI: 10.1021/pr2009302
Keywords
mass spectrometry; protein turnover; N-15; stable isotope labeling; unicellular alga; proteomics; Ostreococcus tauri
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Funding
- Centre for Systems Biology at Edinburgh (CSBE), Centre for Integrative Systems Biology (CISB
- BBSRC
- EPSRC [BB/D019621/1]
- Biotechnology and Biological Sciences Research Council [BB/D019621/1] Funding Source: researchfish
- BBSRC [BB/D019621/1] Funding Source: UKRI
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Protein synthesis and degradation determine the cellular levels of proteins, and their control hence enables organisms to respond to environmental change. Experimentally, these are little known proteome parameters; however, recently, SILAC-based mass cri spectrometry studies have begun to quantify turnover in the proteomes of cell lines, yeast, and animals. Here, we present a proteome-scale method to quantify turnover and calculate synthesis and degradation rate constants of individual proteins in autotrophic organisms such as algae and plants. The workflow is based on the automated analysis of partial stable isotope incorporation with N-15. We applied it in a study of the unicellular pico-alga Ostreococcus tauri and observed high relative turnover in chloroplast-encoded ATPases (0.42-0.58% h(-1)), core photosystem II proteins (0.34-0.51% h(-1)), and RbcL (0.47% h(-1)), while nuclear-encoded RbcS2 is more stable (0.23% h(-1)). Mitochondrial targeted ATPases (0.14-0.16% h(-1)), photosystem antennae (0.09-0.14% h(-1)), and histones (0.07-0.1% h(-1)) were comparatively stable. The calculation of degradation and synthesis rate constants k(deg) and k(syn) confirms RbcL as the bulk contributor to overall protein turnover. This study performed over 144 h of incorporation reveals dynamics of protein complex subunits as well as isoforms targeted to different organelles.
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