Two V-ATPase activity peaks were resolved by sucrose density gradient centrifugation (10–50%, w/w) of the gill microsomal fractions from crabs held in fresh water: a lighter fraction (I) at 24–32% sucrose and a heavier fraction (II) at 39–44% sucrose (Fig. 4), demonstrating that the enzyme occurs in membrane fractions of different densities. Maximum activity was about 2.3-fold greater in peak II, suggesting that the denser membrane fraction is richer in V-ATPase. A broad protein peak, at around 20% sucrose, contained membrane fractions lacking V-ATPase activity.