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Journal Article

Decomposition of time-dependent fluorescence signals reveals codon-specific kinetics of protein synthesis.

MPS-Authors
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Holtkamp,  W.
Department of Physical Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Rodnina,  M. V.
Department of Physical Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Fulltext (public)

2633911.pdf
(Publisher version), 2MB

Supplementary Material (public)

2633911_Suppl_1.htm
(Supplementary material), 238KB

2633911_Suppl_2.pdf
(Supplementary material), 352KB

2633911_Suppl_3.htm
(Supplementary material), 12KB

Citation

Haase, N., Holtkamp, W., Lipowsky, R., Rodnina, M. V., & Rudorf, S. (2018). Decomposition of time-dependent fluorescence signals reveals codon-specific kinetics of protein synthesis. Nucleic Acids Research, 46(22): e130. doi:10.1093/nar/gky740.


Cite as: http://hdl.handle.net/21.11116/0000-0001-F1AB-E
Abstract
During protein synthesis, the nascent peptide chain traverses the peptide exit tunnel of the ribosome. We monitor the co-translational movement of the nascent peptide using a fluorescent probe attached to the N-terminus of the nascent chain. Due to fluorophore quenching, the time-dependent fluorescence signal emitted by an individual peptide is determined by co-translational events, such as secondary structure formation and peptide-tunnel interactions. To obtain information on these individual events, the measured ensemble fluorescence signal has to be decomposed into position-dependent intensities. Here, we describe mRNA translation as a Markov process with specific fluorescence intensities assigned to the different states of the process. Combining the computed stochastic time evolution of the translation process with a sequence of observed ensemble fluorescence time courses, we compute the unknown position-specific intensities and obtain detailed information on the kinetics of the translation process. In particular, we find that translation of poly(U) mRNAs dramatically slows down at the fourth UUU codon. The method presented here detects subtle differences in the position-specific fluorescence intensities and thus provides a novel approach to study translation kinetics in ensemble experiments.