The topology of the proton translocating F0 component of the ATP synthase from E. coli K12: studies with proteases.

The accessibility of the three F0 subunits a, b and c from the Escherichia coli K12 ATP synthase to various proteases was studied in F1‐depleted inverted membrane vesicles. Subunit b was very sensitive to all applied proteases. Chymotrypsin produced a defined fragment of mol. wt. 15,000 which remained tightly bound to the membrane. The cleavage site was located at the C‐terminal region of subunit b. Larger amounts of proteases were necessary to attack subunit a (mol. wt. 30,000). There was no detectable cleavage of subunit c. It is suggested that the major hydrophilic part of subunit b extends from the membrane into the cytoplasm and is in contact with the F1 sector. The F1 sector was found to afford some protection against proteolysis of the b subunit in vitro and in vivo. Protease digestion had no influence on the electro‐impelled H+ conduction via F0 but ATP‐dependent H+ translocation could not be reconstituted upon binding of F1. A possible role for subunit b as a linker between catalytic events on the F1 component and the proton pathway across the membrane is discussed.