Kishimoto T, Kondo H
Exp Cell Res. 1986 Apr;163(2):445-52. doi: 10.1016/0014-4827(86)90075-3.
Cytoplasm of maturing starfish oocytes possesses a factor which induces maturation upon injection into immature oocytes. Such maturation-promoting factor (MPF) was extracted from maturing oocytes of Asterina pectinifera and characterized preliminarily. After 1-methyladenine (1-MeAde) treatment, maturing oocytes were packed in a centrifuge tube to remove jelly and excess medium, and then crushed by centrifugation. The turbid supernatant was homogenized with a buffer containing NaF, Na-beta-glycerophosphate, ATP, EGTA and leupeptin, followed by centrifugation. MPF extracted in the supernatant was purified partially by ammonium sulfate precipitation, hydrophobic chromatography on pentyl-agarose and gel filtration on Sephacryl S-300. The final material induced maturation in the recipient starfish oocytes when 0.5 ng of protein was injected in a volume of 400 pl. The maturation response included germinal vesicle breakdown, and formation of polar bodies and egg pronucleus. Such MPF preparation induced maturation in oocytes of Xenopus laevis as well. Further, starfish MPF was found to be a heat-labile protein; its molecular weight (MW) was estimated as 300 X 10(3) D by gel filtration and its sedimentation coefficient value as 5S by centrifugation on sucrose density gradients.
成熟海星卵母细胞的细胞质中含有一种因子，将其注入未成熟卵母细胞可诱导其成熟。这种成熟促进因子（MPF）是从多棘海盘车成熟卵母细胞中提取并初步鉴定的。用1-甲基腺嘌呤（1-MeAde）处理后，将成熟卵母细胞装入离心管以去除卵胶和多余的培养基，然后通过离心破碎。将浑浊的上清液与含有氟化钠、β-甘油磷酸钠、ATP、乙二醇双四乙酸（EGTA）和亮抑酶肽的缓冲液匀浆，随后进行离心。上清液中提取的MPF通过硫酸铵沉淀、戊基琼脂糖疏水层析和Sephacryl S-300凝胶过滤进行部分纯化。当以400皮升的体积注射0.5纳克蛋白质时，最终产物可诱导受体海星卵母细胞成熟。成熟反应包括生发泡破裂、极体形成和卵原核形成。这种MPF制剂也能诱导非洲爪蟾卵母细胞成熟。此外，发现海星MPF是一种热不稳定蛋白；通过凝胶过滤估计其分子量（MW）为300×10³道尔顿，通过蔗糖密度梯度离心其沉降系数值为5S。
