Secondary structure composition of reconstituted subunit b of the Escherichia coli ATP synthase.
Subunit b of the Escherichia coli ATP synthase was isolated by preparative gel electrophoresis, acetone precipitated and after ion-pair extraction redissolved in a buffer either containing n-dodecyl-beta-D-maltoside or sodium cholate. The secondary structure of isolated subunit b was shown to be the same as within the FO complex, but was strongly dependent on the detergent used for replacement of the phospholipid environment. This was shown by an identical tryptic digestion pattern, which was strongly influenced by the detergent used for solubilization. An influence of the detergent n-dodecyl-beta-D-maltoside on the secondary structure of the hydrophilic part of subunit b was also shown for the soluble part of the polypeptide comprising residues Val25 to Leu156 (bsol) using CD spectroscopy. In order to determine the secondary structure of subunit b in its native conformation, isolated subunit b was reconstituted into E. coli lipid vesicles and analyzed with CD spectroscopy. The resulting spectrum revealed a secondary structure composition of 80% alpha helix together with 14% beta turn conformation. These results suggest that subunit b is not a rigid rod-like alpha helix simply linking F1 to FO, but rather provides an inherent flexibility for the storage of elastic energy within the second stalk generated by rotational movements within the F1FO complex.
Greie
J C
JC
Universität Osnabrück, Fachbereich Biologie/Chemie, Abteilung Mikrobiologie, Germany. greie@biologie.uni-osnabrueck.de
Deckers-Hebestreit
G
G
Altendorf
K
K
D002942:Circular Dichroism; D003902:Detergents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology; D005960:Glucosides / Q000494:pharmacology; D017433:Protein Structure, Secondary; D011510:Proteolipids / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000737:chemistry*; D011522:Protons; D013997:Time Factors; D014357:Trypsin / Q000378:metabolism
D003902:Detergents; D005960:Glucosides; D011510:Proteolipids; D011522:Protons; C020485:proteoliposomes; C040358:dodecyl maltoside; D014357:Trypsin; D006180:Proton-Translocating ATPases