Fig. 3.
DSS cross-linking of NMT2 and ACBD6. Purified hexahistidine-tagged NMT2 (also known as ecNMT2) and ACBD6 recombinant proteins (20 μM each) were incubated in 50 mM potassium phosphate (pH 7.4) at 37°C for 20 min. When indicated, C14-CoA and C18:1-CoA were added at a final concentration of 50 μM. DSS was added to a final concentration of 1 mM and incubated for 30 min at room temperature. Following quenching of DSS with 65 mM Tris-HCl (pH 7.4), proteins were loaded on denaturing SDS-PAGE gels. After electrophoresis, protein bands were detected with Gelcode Blue [(A) and right inset in (B)] or were transferred onto a PVDF membrane and detected with an anti-histidine antibody [left inset in (B)]. The position of the high molecular mass form observed with ACBD6/NMT2 is indicated with an asterisk in the panels. Note the absence of detection of an ACBD6ΔC/NMT2 complex in (B).