Two classes of receptor specific for sperm-activating peptide III in sand-dollar spermatozoa.
We characterized receptors specific for sperm-activating peptide III (SAP-III: DSDSAQNLIQ) in spermatozoa of the sand dollar, Clypeaster japonicus, using both binding and cross-linking techniques. Analyses of the data obtained from the equilibrium binding of a radiolabeled SAP-III analogueto C. japonicus spermatozoa, using Klotz, Scatchard and Hill plots, showed the presence of two classes of receptors specific for SAP-III in the spermatozoa. One of the receptors (high-affinity) had a Kd of 3.4 nM and 3.4 x 10(4) binding sites/spermatozoon. The other receptor (low-affinity) had a Kd of 48 nM, with 6.1 x 10(4) binding sites/spermatozoon. The Kd of the high-affinity receptor was comparable to the median effective concentration of the intracellular-pH-increasing activity of SAP-III and that of the low-affinity receptor was comparable to the median effective concentration of the cellular-cGMP-elevating activity of the peptide. In addition, Scatchard and Hill plots of the data suggested the existence of positive cooperativity between the high-affinity members. Similar results were also obtained from a binding experiment using a sperm-membrane fraction prepared from C. japonicus spermatozoa. The incubation of intact spermatozoa or sperm plasma membranes with the radioiodinated SAP-III analogue and a chemical cross-linking reagent, disuccinimidyl suberate, resulted in the radiolabeling of three proteins with molecular masses of 126, 87 and 64 kDa, estimated by SDS/PAGE under reducing conditions.
Yoshino
K
K
Noto Marine Laboratory, Kanazawa University, Ishikawa, Japan.
Suzuki
N
N
D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D008297:Male; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000378:metabolism*; D012617:Sea Urchins; D013094:Spermatozoa / Q000187:drug effects / Q000378:metabolism*; D013388:Succinimides
D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D009842:Oligopeptides; D011956:Receptors, Cell Surface; D013388:Succinimides; C075201:sperm-activating peptide III, Clypeaster japonicus; C019358:disuccinimidyl suberate