SDS-PAGE and Western Blot Analysis of GvpC Protein

The methods used were those previously described [7 (link), 17 (link)]. Briefly, the SDS-PAGE analysis was performed using the precast Novex^® Tris-glycine gels (4–20%, Invitrogen, Carlsbad, CA, USA). Proteins were then transferred to 0.45 μm nitrocellulose membranes (Millipore Corp., Boston, MA). Membranes were blocked for 30 min in Pierce Fast Blocking Buffer (Thermo Fisher Scientific), incubated in blocking buffer supplemented with Anti-GFP Polyclonal Antibody (Thermo Fisher Scientific) or Anti-GvpC antibodies (GenScript, USA), followed by three washing steps with TBST and then incubation with Goat anti-Rabbit IgG (H + L) Cross-Adsorbed Secondary Antibody, Alexa Fluor 488 (Invitrogen, Catalog # A-11008).