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231226s2023 xx |||||o 00| ||eng c |
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|a 10.1002/adma.202300947
|2 doi
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|a pubmed24n1184.xml
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|a (DE-627)NLM355319675
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|a (NLM)37027309
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|a DE-627
|b ger
|c DE-627
|e rakwb
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| 041 |
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|a eng
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| 100 |
1 |
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|a van Veldhuisen, Thijs W
|e verfasserin
|4 aut
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| 245 |
1 |
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|a Enzymatic Regulation of Protein-Protein Interactions in Artificial Cells
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|c 2023
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|a Text
|b txt
|2 rdacontent
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|a ƒaComputermedien
|b c
|2 rdamedia
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| 338 |
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|a ƒa Online-Ressource
|b cr
|2 rdacarrier
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| 500 |
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|a Date Completed 31.07.2023
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|a Date Revised 31.07.2023
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|a published: Print-Electronic
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|a Citation Status MEDLINE
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|a © 2023 The Authors. Advanced Materials published by Wiley-VCH GmbH.
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|a Membraneless organelles are important for spatial organization of proteins and regulation of intracellular processes. Proteins can be recruited to these condensates by specific protein-protein or protein-nucleic acid interactions, which are often regulated by post-translational modifications. However, the mechanisms behind these dynamic, affinity-based protein recruitment events are not well understood. Here, a coacervate system that incorporates the 14-3-3 scaffold protein to study enzymatically regulated recruitment of 14-3-3-binding proteins is presented, which mostly bind in a phosphorylation-dependent manner. Synthetic coacervates are efficiently loaded with 14-3-3, and phosphorylated binding partners, such as the c-Raf pS233/pS259 peptide (c-Raf), show 14-3-3-dependent sequestration with up to 161-fold increase in local concentration. The c-Raf domain is fused to green fluorescent protein (GFP-c-Raf) to demonstrate recruitment of proteins. In situ phosphorylation of GFP-c-Raf by a kinase leads to enzymatically regulated uptake. The introduction of a phosphatase into coacervates preloaded with the phosphorylated 14-3-3-GFP-c-Raf complex results in a significant cargo efflux mediated by dephosphorylation. Finally, the general applicability of this platform to study protein-protein interactions is demonstrated by the phosphorylation-dependent and 14-3-3-mediated active reconstitution of a split-luciferase inside artificial cells. This work presents an approach to study dynamically regulated protein recruitment in condensates, using native interaction domains
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|a Journal Article
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|a artificial cells
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4 |
|a coacervates
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| 650 |
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4 |
|a phosphorylation
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| 650 |
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4 |
|a scaffold proteins
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| 650 |
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4 |
|a synthetic signaling
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| 650 |
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7 |
|a 14-3-3 Proteins
|2 NLM
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| 650 |
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7 |
|a Peptides
|2 NLM
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| 700 |
1 |
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|a Altenburg, Wiggert J
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Verwiel, Madelief A M
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Lemmens, Lenne J M
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Mason, Alexander F
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Merkx, Maarten
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Brunsveld, Luc
|e verfasserin
|4 aut
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| 700 |
1 |
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|a van Hest, Jan C M
|e verfasserin
|4 aut
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| 773 |
0 |
8 |
|i Enthalten in
|t Advanced materials (Deerfield Beach, Fla.)
|d 1998
|g 35(2023), 29 vom: 05. Juli, Seite e2300947
|w (DE-627)NLM098206397
|x 1521-4095
|7 nnns
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| 773 |
1 |
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|g volume:35
|g year:2023
|g number:29
|g day:05
|g month:07
|g pages:e2300947
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| 856 |
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|u http://dx.doi.org/10.1002/adma.202300947
|3 Volltext
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|a AR
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|d 35
|j 2023
|e 29
|b 05
|c 07
|h e2300947
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