Mammalian spermatozoa contain a membranebound adenylate cyclase which is not sensitive to fluoride, guanine nucleotides, forskolin, or cholera toxin, and which exhibits a striking dependency on Mn’+ relative to Mg2+ for activity [1,2]. Available data suggest that the sperm enzyme lacks a functional guanine nucleotide regulatory component, but may be reconstituted from human erythrocytes [3]. The lack of responsiveness of the sperm adenylate cyclase and the possibility that regulation could be imparted to it, suggested to us that the sperm enzyme may be a useful test system for identifying other components of the adenylate cyclase and for characterizing the interactions of the enzyme’s various subunits. In studies with brain adenylate cyclase analogous to those reported with erythrocytes [3], we found unexpectedly large increases in cyclase activity when preparations from sperm and brain were combined. As we report here, rather than being due to a contribution of a component from brain to the sperm, the superadditive activity was found to be due to a factor in sperm which activated the brain adenylate cyclase. slaughterhouse. Sperms were removed by rinsing the epididymal ducts. After several washes in isotonic buffer, sperms were frozen in small aliquots in liquid nitrogen. Frozen sperm (-10’ cells/ml) were thawed with the addition of 1 or 2 volumes of 10 mM triethanolamine-HCl (pH 7.4) and were homogenized with a motor-driven glassTeflon homogenizer. The particulate material was then collected by centrifugation at 30000 x g for 10 min. The particles were resuspended in 10 mM triethanolamine-HCI (pH 7.4) and were then used for assay of adenylate cyclase. Detergent-dispersed adenylate cyclase from rat brain was prepared as in [4] with the modification that the initial centrifugation speeds were increased from 3000 to 17000 x g. The Lubrol-PX dispersed enzyme was chromatographed on DEAESephadex (A-25) as in [5], with the following modification. Proteins were first eluted with 600 mM NaCl containing 1 mM ethyleneglycol-bis-@aminoethyl ether)N,N’-tetraacetic acid (EGTA) (instead of only 300 mM NaCl). The column was then washed with 600 mM NaCl to wash out the EGTA. The enzyme was then eluted with a solution of 0.25% Lubrol-PX, containing neither salt nor EGTA.