“Structure and Dynamics of the External Stalk of the FoF1-ATP Synthase”

Abstract

The FoF1-ATP synthase is the enzyme responsible for the bulk of ATP synthesized in most organisms. Although the structure and mechanism of the enzyme is generally well-understood, some important intricacies remain unclear. One of the questions still heavily discussed concerns the structure and function of the external stalk which consists of two identical or non-identical subunits b in bacteria and photosynthetic organisms. Making use of structure prediction, de novo modeling, extensive mutagenesis, site-directed spin labeling and ESR spectroscopy, we suggested that the cytosolic, soluble parts of both the E. coli homodimeric b2 as well as a heterodimeric bb’from a cyanobacterial ATP synthase exist as traditional left-handed coiled coils for most of their lengths. Upon binding of the soluble E. coli b2 to soluble F1, the tether region of the b-dimer disengages to form a more loosely packed arrangement that then seems to repack closely before entering the membrane-phase. Using the completely assembled F1Fo-ATPase, we recently observed changes in the packing interactions of b2 during the formation of ATP-hydrolysis transition states. These transitions were observed through the release of spin label catalyzed by non-modified cysteine on the adjacent b-subunit. Only those spin-labels were released that were predicted by us to be close to or at the left-handed coiled coil b-dimer interface, while no release of label from positions that were predicted to be at the outer surface of the helices was observed. The results suggest that while modest conformational changes of the b2-dimer occur during catalysis, the dynamics of these changes do not appear large enough to support a left-handed to right-handed coiled coil conversion that has previously been suggested.