ATP SYNTHASE OF YEAST MITOCHONDRIA - ISOLATION OF THE SUBUNIT-H AND DISRUPTION OF THE ATP14 GENE

A new subunit of the yeast ATP synthase (termed subunit h) has been is olated. Amino acid composition and N-terminal sequencing were determin ed by chemical methods. These data were in agreement with the sequence of the hypothetical protein L8003.20 whose primary structure was dedu ced from DNA sequencing of the yeast chromosome XII. The amino acid se quence encoded by ATP14 gene is 32 amino acids longer than the mature protein, which contains 92 amino acids corresponding to a calculated m ass of 10,408 Da. The protein is hydrophilic and acidic with a calcula ted pH(i) of 4.08. It is not apparently related to any subunit describ ed in other ATP synthases. A null mutant was constructed. The mutation was recessive and the mutant strain was unable to grow on glycerol me dium. A high percentage of rho(-) cells arose spontaneously. The mutan t mitochondria had no detectable oligomycin-sensitive ATPase activity, but still contained ATPase activity with a catalytic sector dissociat ed from the membranous components. The mutant mitochondria did not con tain subunit h, and the mitochondrially encoded hydrophobic subunit 6 was not present.