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bioRxiv · 10.1101/2021.09.22.461445

pSNAP: Proteome-wide analysis of elongating nascent polypeptide chains

Uchiyama, J.·Roy, R.·Wang, D. O.·Yoshino, C.·Mishima, Y.·Ishihama, Y.·Imami, K.
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Abstract

Cellular global translation is often measured using ribosome profiling or quantitative mass spectrometry, but these methods do not provide direct information at the level of elongating nascent polypeptide chains (NPCs) and associated co-translational events. Here we describe pSNAP, a method for proteome-wide profiling of NPCs by affinity enrichment of puromycin- and stable isotope-labeled polypeptides. pSNAP does not require ribosome purification and/or chemical labeling, and captures bona fide NPCs that characteristically exhibit protein N-terminus-biased positions. We applied pSNAP to evaluate the effect of silmitasertib, a potential molecular therapy for cancer, and revealed acute translational repression through casein kinase II and mTOR pathways. We also characterized modifications on NPCs and demonstrated that the combination of different types of modifications, such as acetylation and phosphorylation in the N-terminal region of histone H1.5, can modulate interactions with ribosome-associated factors. Thus, pSNAP provides a framework for dissecting co-translational regulations on a proteome-wide scale.

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Uchiyama, J., Roy, R., Wang, D. O., Yoshino, C., Mishima, Y., Ishihama, Y., Imami, K.. 2021-09-23. pSNAP: Proteome-wide analysis of elongating nascent polypeptide chains. https://doi.org/10.1101/2021.09.22.461445

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