Modulating secretory pathway pH by proton channel co-expression can increase recombinant protein stability in plants.
Eukaryotic expression systems are used for the production of complex secreted proteins. However, recombinant proteins face considerable biochemical challenges along the secretory pathway, including proteolysis and pH variation between organelles. As the use of synthetic biology matures into solutions for protein production, various host-cell engineering approaches are being developed to ameliorate host-cell factors that can limit recombinant protein quality and yield. We report the potential of the influenza M2 ion channel as a novel tool to neutralize the pH in acidic subcellular compartments. Using transient expression in the plant host, Nicotiana benthamiana, we show that ion channel expression can significantly raise pH in the Golgi apparatus and that this can have a strong stabilizing effect on a fusion protein separated by an acid-susceptible linker peptide. We exemplify the utility of this effect in recombinant protein production using influenza hemagglutinin subtypes differentially stable at low pH; the expression of hemagglutinins prone to conformational change in mildly acidic conditions is considerably enhanced by M2 co-expression. The co-expression of a heterologous ion channel to stabilize acid-labile proteins and peptides represents a novel approach to increasing the yield and quality of secreted recombinant proteins in plants and, possibly, in other eukaryotic expression hosts.
Département de phytologie, Université Laval, Québec, Canada.
Philippe V
PV
Jutras
Medicago, Inc., Québec, Canada.
Marc-André
MA
D'Aoust
Medicago, Inc., Québec, Canada.
Manon M-J
MM
Couture
Medicago, Inc., Québec, Canada.
Louis-Philippe
LP
Vézina
Département de phytologie, Université Laval, Québec, Canada.
Marie-Claire
MC
Goulet
Département de phytologie, Université Laval, Québec, Canada. dominique.michaud@fsaa.ulaval.ca.
Dominique
D
Michaud
Département de phytologie, Université Laval, Québec, Canada. f.sainsbury@uq.edu.au.|The University of Queensland, Australian Institute for Bioengineering and Nanotechnology, St Lucia, Australia. f.sainsbury@uq.edu.au.
Frank
F
Sainsbury
D001709:Biotechnology; D006863:Hydrogen-Ion Concentration; D007473:Ion Channels / Q000235:genetics / Q000378:metabolism*; D055550:Protein Stability; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics; D014763:Viral Matrix Proteins / Q000235:genetics / Q000378:metabolism
D007473:Ion Channels; D011994:Recombinant Proteins; D014763:Viral Matrix Proteins
Influenza M2; Intracellular pH; Ion channel expression; Recombinant hemagglutinin; Transient expression