[en] Mussels (Mytilus edulis) adhere to hard surfaces in intertidal marine habitats with a porous underwater glue called the byssus plaque. The plaque is an established role model for bioinspired underwater glues and comprises at least six proteins, most of which are highly cationic and enriched in the post-translationally modified amino acid 3,4-dihydroxyphenylalanine (DOPA). While much is known about the chemistry of plaque adhesion, less is understood about the natural plaque formation process. Here, we investigated plaque structure and formation using 3D electron microscopic imaging, revealing that micro- and nanopores form spontaneously during secretion of protein-filled secretory vesicles. To better understand this process, we developed a method to purify intact secretory vesicles for in vitro assembly studies. We discovered that each vesicle contains a sulfate-associated fluid condensate consisting of ∼9 histidine- and/or DOPA-rich proteins, which are presumably the required ingredients for building a plaque. Rupturing vesicles under specific buffering conditions relevant for natural assembly led to controlled multiphase liquid-liquid phase separation (LLPS) of different proteins, resulting in formation of a continuous phase with coexisting droplets. Rapid coarsening of the droplet phase was arrested through pH-dependent cross-linking of the continuous phase, producing native-like solid porous "microplaques" with droplet proteins remaining as fluid condensates within the pores. Results indicate that histidine deprotonation and sulfates figure prominently in condensate cross-linking. Distilled concepts suggest that combining phase separation with tunable cross-linking kinetics could be effective for microfabricating hierarchically porous materials via self-assembly.
Jehle, Franziska ; Dept. of Chemistry, McGill University, Montreal, Quebec H4A 0B8, Canada ; Dept. of Biomaterials, Max Planck Institute of Colloids and Interfaces, Potsdam 14476, Germany
Duthoo, Emilie ; Université de Mons - UMONS > Faculté des Sciences > Service de Biologie des Organismes Marins et Biomimétisme ; Dept. of Chemistry, McGill University, Montreal, Quebec H4A 0B8, Canada ; Biology of Marine Organisms and Biomimetics Unit, Research Institute for Biosciences, Mons 7000, Belgium
Fratzl, Peter; Dept. of Biomaterials, Max Planck Institute of Colloids and Interfaces, Potsdam 14476, Germany
Bertinetti, Luca ; Dept. of Biomaterials, Max Planck Institute of Colloids and Interfaces, Potsdam 14476, Germany ; B CUBE - Center for Molecular Bioengineering, Technische Universität Dresden, Dresden 01307, Germany
Harrington, Matthew J ; Dept. of Chemistry, McGill University, Montreal, Quebec H4A 0B8, Canada
Language :
English
Title :
Mussels Fabricate Porous Glues via Multiphase Liquid-Liquid Phase Separation of Multiprotein Condensates.
Canada Research Chairs Max-Planck-Gesellschaft Natural Sciences and Engineering Research Council of Canada
Funding text :
We acknowledge A. Wahba and N. Saadeh of the McGill Chemical Characterization (MC) facility for assistance with mass spectrometry. MJH acknowledges funding from the Natural Sciences and Engineering Research Council of Canada Discovery Grant RGPIN-2018-05243 (MJH) and Canada Research Chair Tier 2 Award 950-231953. PF and LB acknowledge support from the Max Planck Society. 2
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