Identification of the dicyclohexylcarbodiimide-reactive protein component of the adenosine 5'-triphosphate energy-transducing system of Escherichia coli.
Membranes of Escherichia coli contain an adenosine 5'-triphosphate (ATP) energy-transducing system that is inhibited by treatment with dicyclohexylcarbodiimide (DCCD). The carbodiimide-reactive protein component of this system has been identified after treatment with [14C]DCCD. This protein has an apparent molecular weight of 9,000 as judged from acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and is extracted from the membrane with chloroform-methanol (2:1). These properties are similar to the analogous protein previously identified in mitochondria (Cattell et al., 1971). A mutant strain, RF-7, has been isolated which derives energy from oxidative phosphorylation in the presence of 5 mM DCCD. The ATP hydrolase activity of the membraned system in the mutant was considerably less sensitive to inhibition by DCCD than that in the wild type. The carbodiimide-reactive protein, which was easily labeled by [14C]DCCD in the wild type, was labeled much less rapidly in the carbodiimide-resistant mutant. It is thus concluded that the reaction of DCCD with this specific protein leads to inhibition of the ATP energy-transducing reactions. The mutation causing carbodiimide resistance in strain RF-7 was mapped. It is cotransduced with the uncA gene at a frequency exceeding 90%. The mutationally altered protein causing the carbodiimide resistance was not conclusively identified. However, reconstitution experiments indicate that the altered protein is not one of the subunits of the soluble ATP hydrolase activity, which can be removed from the membrane by washing with 1 mM tris(hydroxymethyl)aminomethane buffer lacking Mg2+. The carbodiimide-reactive protein remains with the membrane residue after removal of the soluble ATP hydrolase and is thus distinct from these subunits as well.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001426:Bacterial Proteins / Q000032:analysis; D002234:Carbodiimides / Q000494:pharmacology*; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology*; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000032:analysis / Q000187:drug effects / Q000378:metabolism*; D008970:Molecular Weight; D009154:Mutation; D010085:Oxidative Phosphorylation / Q000187:drug effects; D013347:Subcellular Fractions / Q000201:enzymology
D001426:Bacterial Proteins; D002234:Carbodiimides; D004024:Dicyclohexylcarbodiimide; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases