Amino Acid Sequence of the Proteolipid Subunit of the Proton‐Translocating ATPase Complex from the Thermophilic Bacterium PS‐3

Abstract

The proteolipid subunit of the ATPase complex was identified in whole membranes of the thermophilic bacterium PS‐3 by means of a covalent modification with the ¹⁴C‐labelled inhibitor dicyclohexylcarbodiimide. The proteolipid could be purified from the membrane in free and carbodiimide‐modified form by extraction with chloroform/methanol and subsequent carboxymethyl‐cellulose chromatography in mixtures of chloroform/methanol/water. The complete amino acid sequence of the 72‐residue polypeptide could be determined by automated solid‐phase Edman degradation of the whole protein, and of fragments obtained after cleavage with cyanogen bromide and N‐bromosuccinimide. Chemical cleavages and separations of the resulting fragments by gel chromatography were performed in 80% formic acid. The amino acid sequence shows a concentration of hydrophobic amino acids in two segments of about 25 residues at the amino‐terminal and carboxy‐terminal ends. The polar residues are clustered in the middle of the polypeptide chain. The bound [¹⁴C]dicyclohexylcarbodiimide label is recovered exclusively at position 56, which is occupied by a glutamyl residue. The proteolipid from PS‐3 exhibits homology to the corresponding ATPase subunit from mitochondria. The carbodiimide‐reactive glutamyl residue occurs at the position as in the mitochondrial proteins.