The catalytic subunit of a membrane-bound pyrophosphatase was purified by electroendosmotic preparative
electrophoresis from etiolated pea stem mitochondria. The enzyme was identified as a single
peak relatively pure, because only a very limited number of polypeptides were detectable by SDSlPAGE
of the active fractions. The pyrophosphatase was associated to a band with a molecular mass of 35 kDa,
showing a specific activity of 0.7 pmol P, ' mg-' protein . min-' (37"C, pH 8.0) and an apparent
K,,, value of 200 pM. The hydrolytic activity required Mgz+, was inhibited by imidodiphosphate
(HNO,P,Na,), Ca", F- and was stimulated by phospholipids. Cardiolipin, phophatidylcholine and
phosphatidylethanolamine had the maximal activating effect.
The isolated protein is very similar to the catalytic subunit of pyrophosphatases isolated from rat liver
@-subunit) and Saccharomyces cerevisiae mitochondria.
