Can the b fusion protein, replace b and  subunits in E.coli?

ATP synthase is a ubiquitous membrane enzyme that plays a key role in biological energy metabolism. In Escherichia coli, the simplest form contains eight different subunits, with a stoichiometry of α₃β₃γδεab₂c₁₀. ATP synthase uses a unique rotational mechanism; γεc₁₀ form the rotor and α₃β₃δab₂ the stator. The stator stalk which is composed of two copies of subunit b and one copy of subunit δ is the least well understood part of ATP synthase in terms of structure. To understand the energetic contributions of the individual b subunits to binding  to F_1, construction of a fusion protein between one of the b subunits and δ was the main intention of the project. Such a fusion protein does exist in Mycobacteria. Initially the bδ fusion protein as a part of the ATP synthase holoenzyme was expressed, using plasmid pbwu13.4. The intergenic region was replaced by a synthetic DNA fragment encoding the connecting domain of the bδ fusion protein of Mycobacterium vanbaalenii. As the final step in the construction of the expression vector another copy of gene encoding the b subunit was inserted before the gene encoding for the fusion protein. After transformation into strain DK8, these strains showed growth on limiting glucose concentration and succinate plates confirming that these strains still have the ATP synthase activity. The isolated membranes form the strains carrying the fusion protein showed ATPase acitivity. Therefore the functional assays confirm that this fusion protein can replace b and δ of E.coli.