Bacterial Expression and Purification of Protein Complexes

The bsDbs were prepared by using the bacterial expression system we described previously^{13 (link)}. Briefly, the constructs were expressed individually in E. coli strain BL21 Star (DE3) (Life Technologies, Carlsbad, CA, USA) and purified from the bacterial supernatant and periplasmic fractions. The constructs were also prepared from the intracellular soluble fraction by using BugBuster reagent (Merck KGaA, Darmstadt, Germany) according to the manufacturer’s instructions^{34 (link)}. After purification by immobilized metal-affinity chromatography, gel filtration analysis (HiLoad Superdex 200-pg column 26/60, GE Healthcare Bio-Science, Piscataway, NJ, USA) was used to fractionate the dimers of each bsDb and tetramers of LH-HY52W. The column was equilibrated with PBS, and then purified bsDb was loaded onto the column at a flow rate of 2–2.5 mL/min.