Schaller J, Akiyama K, Tsuda R, Hara M, Marti T, Rickli E E
Institute of Biochemistry, University of Bern, Switzerland.
Eur J Biochem. 1987 Dec 30;170(1-2):111-20. doi: 10.1111/j.1432-1033.1987.tb13674.x.
gamma-Seminoprotein (gamma-SM), a glycoprotein from human seminal plasma, was isolated in highly purified form by ion-exchange chromatography on a Mono Q column. The main form, fraction M, was homogeneous by PAGE at pH 8.3 and by SDS-PAGE. The complete amino acid sequence of gamma-SM was determined with the aid of fragments generated by cleavages with cyanogen bromide, clostripain, chymotrypsin and Staphylococcus aureus V8 protease. The fragments were aligned with overlapping sequences. The single polypeptide chain of gamma-SM contains 237 amino acids with a calculated Mr of 26079. A single N-linked carbohydrate side chain is attached to Asn45. The complex structure of this oligosaccharide has been determined recently [van Halbeek H. et al. (1985) Biochem. Biophys. Res. Commun. 131, 507-514]. Sequence comparison with serine proteases shows a high degree of homology, especially with the kallikrein family. The residues in the vicinity of the active site of serine proteases are also highly conserved in gamma-SM, indicating the participation of His41, Asp96 and Ser189 in its active site. gamma-SM hydrolyzed M-casein with a pH optimum at 8.0, but failed to hydrolyze any of the synthetic substrates tested. This proteolytic activity could be inhibited with diisopropylfluorophosphate, phenylmethylsulfonyl fluoride, Zn2+ or Hg2+ ions.
γ-精浆蛋白（γ-SM）是一种来自人类精浆的糖蛋白，通过在Mono Q柱上进行离子交换色谱法以高纯度形式分离得到。主要形式的组分M，在pH 8.3的聚丙烯酰胺凝胶电泳（PAGE）和十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）中均显示为均一的。借助溴化氰、梭菌蛋白酶、胰凝乳蛋白酶和金黄色葡萄球菌V8蛋白酶切割产生的片段，确定了γ-SM的完整氨基酸序列。这些片段通过重叠序列进行比对。γ-SM的单条多肽链包含237个氨基酸，计算得出的分子量为26079。一个N-连接的碳水化合物侧链连接到Asn45上。这种寡糖的复杂结构最近已被确定[van Halbeek H.等人（1985年）《生物化学与生物物理研究通讯》131, 507 - 514]。与丝氨酸蛋白酶的序列比较显示出高度同源性，尤其是与激肽释放酶家族。丝氨酸蛋白酶活性位点附近的残基在γ-SM中也高度保守，表明His41、Asp96和Ser189参与其活性位点。γ-SM在pH最适值为8.0时水解M-酪蛋白，但未能水解所测试的任何一种合成底物。这种蛋白水解活性可被二异丙基氟磷酸酯、苯甲基磺酰氟、Zn2 +或Hg2 +离子抑制。
