Increased yields and biological potency of knob-into-hole-based soluble MHC class II molecules

Pau Serra (), Nahir Garabatos, Santiswarup Singha, César Fandos, Josep Garnica, Patricia Solé, Daniel Parras, Jun Yamanouchi, Jesús Blanco, Meritxell Tort, Mireia Ortega, Yang Yang, Kristofor K. Ellestad and Pere Santamaria ()
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Pau Serra: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Nahir Garabatos: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Santiswarup Singha: University of Calgary
César Fandos: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Josep Garnica: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Patricia Solé: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Daniel Parras: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Jun Yamanouchi: University of Calgary
Jesús Blanco: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Meritxell Tort: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Mireia Ortega: Institut D’Investigacions Biomèdiques August Pi i Sunyer
Yang Yang: University of Calgary
Kristofor K. Ellestad: University of Calgary
Pere Santamaria: Institut D’Investigacions Biomèdiques August Pi i Sunyer

Nature Communications, 2019, vol. 10, issue 1, 1-14

Abstract: Abstract Assembly of soluble peptide-major histocompatibility complex class II (pMHCII) monomers into multimeric structures enables the detection of antigen-specific CD4+ T cells in biological samples and, in some configurations, their reprogramming in vivo. Unfortunately, current MHCII-αβ chain heterodimerization strategies are typically associated with low production yields and require the use of foreign affinity tags for purification, precluding therapeutic applications in humans. Here, we show that fusion of peptide-tethered or empty MHCII-αβ chains to the IgG1-Fc mutated to form knob-into-hole structures results in the assembly of highly stable pMHCII monomers. This design enables the expression and rapid purification of challenging pMHCII types at high yields without the need for leucine zippers and purification affinity tags. Importantly, this design increases the antigen-receptor signaling potency of multimerized derivatives useful for therapeutic applications and facilitates the detection and amplification of low-avidity T cell specificities in biological samples using flow cytometry.

Date: 2019
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DOI: 10.1038/s41467-019-12902-2

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