A novel microfluidics-based method for probing weak protein-protein interactions.
We report the use of a novel microfluidics-based method to detect weak protein-protein interactions between membrane proteins. The tight junction protein, claudin-2, synthesised in vitro using a cell-free expression system in the presence of polymer vesicles as membrane scaffolds, was used as a model membrane protein. Individual claudin-2 molecules interact weakly, although the cumulative effect of these interactions is significant. This effect results in a transient decrease of average vesicle dispersivity and reduction in transport speed of claudin-2-functionalised vesicles. Polymer vesicles functionalised with claudin-2 were perfused through a microfluidic channel and the time taken to traverse a defined distance within the channel was measured. Functionalised vesicles took 1.19 to 1.69 times longer to traverse this distance than unfunctionalised ones. Coating the channel walls with protein A and incubating the vesicles with anti-claudin-2 antibodies prior to perfusion resulted in the functionalised vesicles taking 1.75 to 2.5 times longer to traverse this distance compared to the controls. The data show that our system is able to detect weak as well as strong protein-protein interactions. This system offers researchers a portable, easily operated and customizable platform for the study of weak protein-protein interactions, particularly between membrane proteins.
Institute of Materials Research and Engineering, 3 Research Link, Singapore. tancwd@imre.a-star.edu.sg
Darren Cherng-wen
DC
Tan
I Putu Mahendra
IP
Wijaya
Mirjam
M
Andreasson-Ochsner
Elena Nikolaevna
EN
Vasina
Madhavan
M
Nallani
Walter
W
Hunziker
Eva-Kathrin
EK
Sinner
D000906:Antibodies / Q000276:immunology; D057167:Claudins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D004867:Equipment Design; D017022:Flow Injection Analysis; D006801:Humans; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D011108:Polymers / Q000737:chemistry; D025941:Protein Interaction Mapping / Q000295:instrumentation*; D013205:Staphylococcal Protein A / Q000737:chemistry; D013211:Staphylococcus aureus / Q000737:chemistry
D000906:Antibodies; C491875:CLDN2 protein, human; D057167:Claudins; D011108:Polymers; D013205:Staphylococcal Protein A