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  Quantitative phase microscopy enables precise and efficient determination of biomolecular condensate composition

McCall, P. M., Kim, K., Fritsch, A. W., Iglesias-Artola, J., Jawerth, L., Wang, J., et al. (2020). Quantitative phase microscopy enables precise and efficient determination of biomolecular condensate composition. bioRxiv. doi:10.1101/2020.10.25.352823.

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2020.10.25.352823v1.full.pdf (Preprint), 13MB
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2020.10.25.352823v1.full.pdf
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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-ND 4.0 International license.

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McCall, Patrick M.1, Author
Kim, Kyoohyun2, 3, Author              
Fritsch, Anatol W.1, Author
Iglesias-Artola, J.M.1, Author
Jawerth, L.M.1, Author
Wang, Jie1, Author
Ruer, M.1, Author
Peychl, J.1, Author
Poznyakovskiy, Andrey1, Author
Guck, Jochen2, 3, Author              
Alberti, Simon1, Author
Hyman, Anthony A.1, Author
Brugués, Jan1, Author
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1external, ou_persistent22              
2Guck Division, Max Planck Institute for the Science of Light, Max Planck Society, ou_3164416              
3Technische Universität Dresden, ou_persistent22              

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 Abstract: Many compartments in eukaryotic cells are protein-rich biomolecular condensates demixed from the cyto- or nucleoplasm. Although much has been learned in recent years about the integral roles condensates play in many cellular processes as well as the biophysical properties of reconstituted condensates, an understanding of their most basic feature, their composition, remains elusive. Here we combined quantitative phase microscopy (QPM) and the physics of sessile droplets to develop a precise method to measure the shape and composition of individual model condensates. This technique does not rely on fluorescent dyes or tags, which we show can significantly alter protein phase behavior, and requires 1000-fold less material than traditional label-free technologies. We further show that this QPM method measures the protein concentration in condensates to a 3-fold higher precision than the next best label-free approach, and that commonly employed strategies based on fluorescence intensity dramatically underestimate these concentrations by as much as 50-fold. Interestingly, we find that condensed-phase protein concentrations can span a broad range, with PGL3, TAF15(RBD) and FUS condensates falling between 80 and 500 mg/ml under typical in vitro conditions. This points to a natural diversity in condensate composition specified by protein sequence. We were also able to measure temperature-dependent phase equilibria with QPM, an essential step towards relating phase behavior to the underlying physics and chemistry. Finally, time-resolved QPM reveals that PGL3 condensates undergo a contraction-like process during aging which leads to doubling of the internal protein concentration coupled to condensate shrinkage. We anticipate that this new approach will enable understanding the physical properties of biomolecular condensates and their function.

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Language(s): eng - English
 Dates: 2020-10-25
 Publication Status: Published online
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 Identifiers: DOI: 10.1101/2020.10.25.352823
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Title: bioRxiv
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