Presentation Information
[P04-499]An integrated approach for assessing state-specific protein turnover via molecular weight fractionation and isotopic labeling
○Tokichiro Abe1, Atsushi Hatano1, Masaki Matsumoto1 (1. Niigata Univ. (Japan))
Keywords:
Proteomics,system biology,protein turnover,SEC,pSILAC
[Purpose]
To elucidate the regulatory principles governing protein turnover at the proteome level, with a particular focus on how protein complex formation influences protein degradation dynamics. While previous studies have suggested that non-exponential decay is frequently observed among proteins that form complexes, the extent to which complex formation directly modulates degradation rates remains unclear. This study aims to systematically investigate the impact of protein–protein interactions within complexes on protein turnover.
[Method]
To achieve this, we developed an integrated experimental approach that combines size exclusion chromatography coupled with mass spectrometry (SEC-MS) and pulsed stable isotope labeling by amino acids in cell culture (pSILAC). This approach enables the simultaneous characterization of protein complex states and the measurement of protein turnover dynamics. By applying this method at the proteome scale, we performed a comprehensive and quantitative analysis to assess how complex formation influences protein degradation behavior.
[Results]
Integration of pSILAC-SEC-MS data revealed that protein turnover rates were broadly distributed across SEC fractions. While no significant differences were observed when fractions were grouped solely by molecular weight, proteins belonging to large multimeric complexes exhibited lower turnover rates in high-molecular-weight fractions and higher turnover rates in low-molecular-weight fractions. Furthermore, under growth arrest conditions, proteins annotated as complex subunits showed a significant reduction in turnover rates compared to non-annotated proteins, indicating that protein stability is jointly influenced by complex formation and cellular proliferation.
[Consideration]
Previous studies suggest that many proteins follow an exponential decay model, whereas proteins within complexes often exhibit non-exponential behavior. Our results further indicate that protein stability is strongly dependent on assembly state, with multimeric complex subunits being stabilized in high-molecular-weight fractions and destabilized in monomeric states. Moreover, the reduction in turnover rates under growth-arrest conditions suggests that dilution through cell division is a major determinant of apparent turnover for stable complexes.
[Conclusion]
This study establishes an integrated and quantitative framework to evaluate the impact of protein complex formation on protein turnover at the proteome scale. Our findings demonstrate that protein stability is not solely determined by intrinsic sequence features, but is dynamically regulated by assembly state and cellular context, including proliferation. These insights provide a new perspective on how cells control protein abundance and maintain homeostasis through the coordinated regulation of synthesis, degradation, and complex formation.
To elucidate the regulatory principles governing protein turnover at the proteome level, with a particular focus on how protein complex formation influences protein degradation dynamics. While previous studies have suggested that non-exponential decay is frequently observed among proteins that form complexes, the extent to which complex formation directly modulates degradation rates remains unclear. This study aims to systematically investigate the impact of protein–protein interactions within complexes on protein turnover.
[Method]
To achieve this, we developed an integrated experimental approach that combines size exclusion chromatography coupled with mass spectrometry (SEC-MS) and pulsed stable isotope labeling by amino acids in cell culture (pSILAC). This approach enables the simultaneous characterization of protein complex states and the measurement of protein turnover dynamics. By applying this method at the proteome scale, we performed a comprehensive and quantitative analysis to assess how complex formation influences protein degradation behavior.
[Results]
Integration of pSILAC-SEC-MS data revealed that protein turnover rates were broadly distributed across SEC fractions. While no significant differences were observed when fractions were grouped solely by molecular weight, proteins belonging to large multimeric complexes exhibited lower turnover rates in high-molecular-weight fractions and higher turnover rates in low-molecular-weight fractions. Furthermore, under growth arrest conditions, proteins annotated as complex subunits showed a significant reduction in turnover rates compared to non-annotated proteins, indicating that protein stability is jointly influenced by complex formation and cellular proliferation.
[Consideration]
Previous studies suggest that many proteins follow an exponential decay model, whereas proteins within complexes often exhibit non-exponential behavior. Our results further indicate that protein stability is strongly dependent on assembly state, with multimeric complex subunits being stabilized in high-molecular-weight fractions and destabilized in monomeric states. Moreover, the reduction in turnover rates under growth-arrest conditions suggests that dilution through cell division is a major determinant of apparent turnover for stable complexes.
[Conclusion]
This study establishes an integrated and quantitative framework to evaluate the impact of protein complex formation on protein turnover at the proteome scale. Our findings demonstrate that protein stability is not solely determined by intrinsic sequence features, but is dynamically regulated by assembly state and cellular context, including proliferation. These insights provide a new perspective on how cells control protein abundance and maintain homeostasis through the coordinated regulation of synthesis, degradation, and complex formation.
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