Measuring the dynamics of E. coli ribosome biogenesis using pulse-labeling and quantitative mass spectrometryElectronic supplementary information (ESI) available. See DOI: 10.1039/c2mb25310k
The ribosome is an essential organelle responsible for cellular protein synthesis. Until recently, the study of ribosome assembly has been largely limited to in vitro assays, with few attempts to reconcile these results with the more complex ribosome biogenesis process inside the living cell. Here,...
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Main Authors | , , , , |
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Format | Journal Article |
Language | English |
Published |
30.10.2012
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Online Access | Get full text |
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Summary: | The ribosome is an essential organelle responsible for cellular protein synthesis. Until recently, the study of ribosome assembly has been largely limited to
in vitro
assays, with few attempts to reconcile these results with the more complex ribosome biogenesis process inside the living cell. Here, we characterize the ribosome synthesis and assembly pathway for each of the
E. coli
ribosomal protein (r-protein)
in vivo
using a stable isotope pulse-labeling timecourse. Isotope incorporation into assembled ribosomes was measured by quantitative mass spectrometry (qMS) and fit using steady-state flux models. Most r-proteins exhibit precursor pools ranging in size from 0% to 7% of completed ribosomes, and the sizes of these individual r-protein pools correlate well with the order of r-protein binding
in vitro.
Additionally, we observe anomalously large precursor pools for specific r-proteins with known extra-ribosomal functions, as well as three r-proteins that apparently turnover during steady-state growth. Taken together, this highly precise, time-dependent proteomic qMS approach should prove useful in future studies of ribosome biogenesis and could be easily extended to explore other complex biological processes in a cellular context.
We developed a high-precision stable isotope pulse-labeling approach to quantitate key features of ribosome biogenesis in
E. coli
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Bibliography: | 10.1039/c2mb25310k Electronic supplementary information (ESI) available. See DOI |
ISSN: | 1742-206X 1742-2051 |
DOI: | 10.1039/c2mb25310k |