Purification and characterization of cleavage accelerating substances extracted with acid sea water from unfertilized sea urchin eggs

从未受精的海胆卵中提取的酸性海水提取物（卵提取物：EE）可以诱导海胆卵的人工活化、加速卵裂和动物化。在这项研究中，我们试图纯化和表征卵裂加速因子（CAF）在EE。EE至少包含两种加速因素；即胶袋可过滤型（MW < 12,000）和胶袋不可过滤型（MW <大于等于>12,000）。首先，我们尝试提纯低分子量的前一因子。采用凝胶过滤（Bio gel P-6和P-2）和高效液相色谱（ODS C-18）对胶袋过滤因子进行浓缩和分离。因此，我们可以得到一个非常有效的CAF，它似乎是一个核苷酸或核苷（MW: < <）。另一方面，将胶袋-非过滤因子应用于DEAE Sephadex a -50，得到一个含有CAF的分数。在这个部分中，检测到少量的蛋白质，因此该因子似乎是核苷酸或核苷，结合了一些蛋白质。此外，还发现胶袋不可过滤因子中含有一种能完全阻止细胞分裂的卵裂抑制因子。因此，细胞周期似乎是由这两种因子的对跖作用的平衡来调节的，尽管这种抑制因子还需要更精确的研究。结果表明，e - e处理后，细胞质游离Ca^+浓度和细胞质pH均未升高，但磷酸精氨酸含量升高至受精卵水平。这种变化可能与电子离子处理引起的解理加速有关。

The acid sea water extract from unfertilized sea urchin eggs (egg extract:EE) can induce artificial activation, acceleration of cleavage, and animalization in sea urchin eggs. In this study, we attempted to purify and characterize cleavage accelerating factors (CAF) in EE.EE contains at least two kinds of accelerating factors; that is, a collodion bag-filterable one(MW < 12,000) and a collodion bag-non-filterable one (MW <greater than or equal>12,000). At first, we have tried to purify the former factor with low molecular weight. This collodion bag filterable factor was concentrated and fractionated by gel filtration (Bio Gel P-6 and P-2) and HPLC (ODS C-18). Thus we could obtained a very effective fraction of CAF which seemed to be a nucleotide or nuleoside (MW: <less thanOn the other hand, the collodion bag-non-filterable factor was applied to DEAE Sephadex A-50 and a fraction containing CAF was obtained. In this fraction, a small amount of proteins was detected, so that the factor seemed to be a nucleotide or nucleoside, binding some proteins.Furthermore, it was found that the collodion bag-non- filterable factor contains a cleavage inhibiting factor which stops cytokinesis completely. Therefore, it seems that a cell cycle is regulated by balance of two antipodal actions of such factors, although this inhibiting factor should be studied more precisely.It was also revealed that neither cytoplasmic free Ca^+ concentration nor cytoplasmic pH are not raised by EE-treatment, but the content of arginine phosphate is increased to the level in fertilized eggs. Such a changes may have some relationship to acceleration of cleavage induced by EE-treatment.