Mode of action of sperm-activating peptide

在过去的十年里，我们从分布在5个分类目的17种海胆的卵冻中纯化了74种精子激活肽。这些多肽对海胆精子的生物学效应基本相同，尽管它们的生物效应和结构在序数水平上是特异性的。根据这种特异性，这些肽可以分为五类。精子激活肽I （sp -I: GFDLNGGGVG）特异性交联到一个71 kDa的蛋白上，并诱导了半心鼠精子质膜鸟苷酸环化酶的电泳迁移率变化，同时该酶的去磷酸化和失活。鸟苷酸环化酶的去磷酸化和随后的失活在花青素a的存在下受到抑制。磷酸化和去磷酸化形式的鸟苷酸环化酶每摩尔蛋白质分别含有约24和4摩尔磷酸。从雄性毛鼠cDNA文库中分离出了一个谷丁酸环化酶cDNA克隆。开放阅读框预测1125个氨基酸的蛋白质，包括21个残基的表观信号肽；一个由25个氨基酸组成的单跨膜结构域将成熟的蛋白质分成由485个氨基酸组成的氨基端胞外结构域和由594个细胞内氨基酸组成的羧基结构域。胞内结构域具有激酶样区和催化区。在类激酶区和催化区分别存在9个和15个丝氨酸残基。我们从精子尾部纯化了三种不同的蛋白磷酸酶。calyculin A抑制了这些酶的活性。

In the past decade, we purified 74 sperm-activating peptides from the egg jelly of 17 species of sea urchins distributed over five taxonomic orders. These peptides show essentially the same biological effects on sea urchin spermatozoa although the biological effects and structures of the peptides are specific at the ordinal level. With regard to that specificity, these peptides can be classified into five groups. Sperm-activating peptide I (SAP-I : GFDLNGGGVG) crosslinked specifically to a 71 kDa protein and induced an electrophoretic mobility change in the guanylate cyclase of Hemicentrotus pulcherrimus sperm plasma membrane with concomitant dephosphorylation and inactivation of the enzyme. The dephosphorylation and subsequent inactivation of the enzyme were inhibited in the presence of calyculin A. The phosphorylated and dephosphorylated forms of the guanylate cyclase contained about 24 and 4 moles of phosphate per mol protein, respectively. A cDNA clone for the gudnylate cyclase has been isolated from a cDNA library for H. pulcherrimus testes. An open reading frame predicted a protein of 1125 amino acids including an apparent signal peptide of 21 residues ; a single transmembrane domain of 25 amino acids divided the mature protein into an amino-terminal, extracellular domain of 485 amino acids and carboxyl domain of 594 intracellular amino acids. The intracellular domain possessed a kinase-like region and a catalytic region. Nine and fifteen serine residues existed in the kinase-like and the catalytic regions, respectively. We purified three different protein phosphatases from the sperm tails. The activity of these enzyme was inhibited by calyculin A.