Coupling artificial actin cortices to biofunctionalized lipid monolayers.
We report the assembly of protein supramolecular structures at an air-water interface and coupling of artificial actin cortices to such structures. The coupling strategies adopted include electrostatic binding of actin to monolayers doped with lipids, exposing positively charged poly(ethylene glycol) headgroups; binding of biotinylated actin to lipids carrying biotin headgroups through avidin; binding of actin to membranes through biotinylated hisactophilin (a cellular actin-membrane coupler) using an avidin-biotin linkage; and coupling of actin to membranes carrying chelating lipids through a 15-nm-diameter protein capsid (bacterial lumazine synthase or LuSy) exhibiting histidine tags (which bind both to actin and to the chelating lipid). The distribution of the proteins in a direction normal to the interface was measured by neutron reflectivity under different conditions of pH and ionic strength. In the case of the first three binding methods, the thickness of the actin film was found to correspond to a single actin filament. Multilayers of actin could be formed only by using the multifunctional LuSy couplers that exhibit 60 hexahistidine tags and can thus act as actin cross-linkers. The LuSy-mediated binding can be reversibly switched by pH variations.
Sengupta
Kheya
K
Lehrstuhl für Biophysik, E22, Technische Universität München, James-Franck-Strasse 1, D-85748 Garching, Germany. k.sengupta@fz-juelich.de
Limozin
Laurent
L
Tristl
Matthias
M
Haase
Ilka
I
Fischer
Markus
M
Sackmann
Erich
E
D000199:Actins / Q000737:chemistry*; D000818:Animals; D001665:Binding Sites; D066298:In Vitro Techniques; D008051:Lipid Bilayers / Q000737:chemistry; D008055:Lipids / Q000737:chemistry*; D008840:Microfilament Proteins / Q000737:chemistry; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry; D046912:Multiprotein Complexes / Q000737:chemistry; D009502:Neutrons; D011485:Protein Binding; D015800:Protozoan Proteins / Q000737:chemistry; D055672:Static Electricity; D013499:Surface Properties
D000199:Actins; D008051:Lipid Bilayers; D008055:Lipids; D008840:Microfilament Proteins; D009097:Multienzyme Complexes; D046912:Multiprotein Complexes; D015800:Protozoan Proteins; C058561:hisactophilin protein, Protozoan; C044822:6,7-dimethyl-8-ribityllumazine synthase