The structure of bovine F-1-ATPase complexed with the antibiotic inhibitor aurovertin B

In the structure of bovine mitochondrial F-1-ATPase that was previously determined with crystals grown in the presence of adenylyl-imidodiphosphate (AMP-PNP) and ADP, the three catalytic beta-subunits have different conformations and nucleotide occupancies. Adenyl-imidodiphosphate is bound to one beta-subunit (beta(TP)), ADP is bound to the second (beta(DP)), and no nucleotide is bound to the third (beta(E)). Here we show that the uncompetitive inhibitor aurovertin B binds to bovine F-1 at two equivalent sites in beta(TP) and beta(E), in a cleft between the nucleotide binding and C-terminal domains. In beta(DP), the aurovertin B pocket is incomplete and is inaccessible to the inhibitor. The aurovertin B bound to beta(TP) interacts with alpha-Glu399 in the adjacent alpha(TP) subunit, whereas the aurovertin B bound to beta(E) is too distant from alpha(E) to make an equivalent interaction. Both sites encompass beta Arg-412, which was shown by mutational studies to be involved in binding aurovertin. Except for minor changes around the aurovertin pockets, the structure of bovine F-1-ATPase is the same as determined previously. Aurovertin B appears to act by preventing closure of the catalytic interfaces, which is essential for a catalytic mechanism involving cyclic interconversion of catalytic sites.