Comparative analysis of proteins from the fibrous sheath and outer dense fibers of rat spermatozoa.
Comparative analysis of proteins from the fibrous sheath and outer dense fibers of rat spermatozoa.
复制标题
大鼠精子纤维鞘和外致密纤维蛋白质的比较分析。
DOI：
10.1095/biolreprod39.1.169
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发表时间：
1988
影响因子：
3.6
通讯作者：
R. Oko
中科院分区：
文献类型：
--
作者：
R. Oko
采用聚丙烯酰胺凝胶电泳和免疫化学方法，对精子尾部两种细胞骨架成分纤维鞘（FS）和外致密纤维（ODF）的蛋白质组成进行了比较。分离的FS和ODF，其纯度通过电子显微镜（EM）验证，变性，或运行在十二烷基硫酸钠-聚丙烯酰胺凝胶或用于提高抗体。凝胶分别显示了至少18条和14条针对FS和ODF的多肽带。FS主要条带的分子质量分别为75、27.5和14.4 kDa， odf连接段主要条带的分子质量分别为32-26、20、14.4、84和80 kDa。在凝胶上发现了几个突出的FS和ODF条带，14.4 kDa多肽具有相似的电泳特性。抗FS血清与大多数Western blot转移的FS多肽发生反应，但也与14.4 kDa的ODF多肽发生强烈交叉反应，与其他主要ODF多肽的亲和力较低。抗ODF血清与大多数ODF多肽发生反应，但也与14.4 kDa的主要FS多肽发生强烈交叉反应，而与其他几个FS多肽（包括75 kDa带）的亲和力较低。从14.4 kDa FS多肽中亲和纯化的抗体仅与14.4 kDa ODF多肽交叉反应，而从14.4 kDa ODF多肽中纯化的抗体与14.4、27.5、57和63 kDa FS多肽交叉反应。免疫细胞化学方法应用于精子，证实了免疫印迹观察到的免疫交叉反应。这项研究表明，FS和ODF都是由许多多肽组成的，其中一些具有相似的分子量，它们是相关的细胞骨架结构，因为它们具有共同的表位，并且都含有14.4 kDa的多肽，具有共同的抗原和电泳特性。
The protein composition of the fibrous sheath (FS) and the outer dense fibers (ODF), two cytoskeletal components of the tail of spermatozoa, was compared by using polyacrylamide gel electrophoresis and immunochemistry applied to Western blots and to spermatozoa. Isolated FS and ODF, the purity of which were verified by electron microscopy (EM), were denatured and either run on sodium dodecyl sulfate-polyacrylamide gels or used to raise antibodies. The gels revealed at least 18 and 14 polypeptide bands for the FS and ODF, respectively. The major bands of the FS had molecular masses of 75, 27.5, and 14.4 kDa, whereas the major bands of the ODF-connecting piece had molecular masses of 32-26, 20, 14.4, 84, and 80 kDa. Several prominent FS and ODF bands were found to comigrate on gels, and the 14.4 kDa polypeptides had similar electrophoretic properties. Anti-FS serum reacted with the majority of Western blot-transferred FS polypeptides, but also cross-reacted strongly with a major 14.4 kDa ODF polypeptide and with less affinity to other major ODF polypeptides. Anti-ODF serum reacted with the majority of ODF polypeptides, but also cross-reacted strongly with a major 14.4 kDa FS polypeptide, and with less affinity to several other FS polypeptides including the 75 kDa band. Antibodies affinity-purified from the 14.4 kDa FS polypeptide only cross-reacted with the 14.4 kDa ODF polypeptide, whereas antibodies purified from the 14.4 kDa ODF polypeptide cross-reacted with 14.4, 27.5, 57, and 63 kDa FS polypeptides. The immunocross-reactions observed on Western blots were confirmed by immunocytochemical methods applied to spermatozoa. This study demonstrates that the FS and ODF, both composed of many polypeptides, several having similar molecular weights, are related cytoskeletal structures as they have epitopes in common, and both contain 14.4 kDa polypeptides with common antigenic and electrophoretic properties.