Vanadium ion inhibition of alkaline phosphatase-catalyzed phosphate ester hydrolysis.

The intracellular location of guanylate cyclase was examined in sperm from two species of sea urchin, Strongylocentrotus purpuratus and Lytechinus pictus, and from the tube worm Chaetopterus variopedatus. Cells suspended in a medium isotonic with sea water were passed repeatedly through a 23-gauge hypodermic needle to break flagella from heads. This preparation was then fractionated by two methods, one based on centrifugation over a 25% sucrose medium and the other involving repeated differential centrifugation, to resolve flagella from heads. Guanylate cyclase specific activity was increased 3.5–4.5-fold in the flagellar fraction relative to the starting sperm homogenate. Relatively little activity was present in the head fraction where specific activity was $\text{1}\text{10}\text{–}\text{1}\text{100}$ that of the flagella. Plasma membranes were separated from axonemal microtubules by dialyzing flagella against low ionic strength buffer, followed by centrifugation over a 40% sucrose medium. Although the overall recovery of guanylate cyclase was low, the specific activity in the plasma membrane fraction was increased two- to threefold over the dialyzed flagella, and over 90% of the recovered activity resided in this fraction. Thus the flagellar plasma membrane is a site rich in guanylate cyclase. It could not be determined, however, whether this is the only intracellular locale of the enzyme.