人睾丸蛋白NYD-SP16的功能研究

发布时间：2018-10-25 14:06

【摘要】：本实验室前期的工作中，在人睾丸cDNA microarray差异杂交的基础上，克隆了一个新的成人睾丸高表达的基因，命名为NYD-SP16。序列分析显示NYD-SP16与小鼠睾丸基因AK006554的核苷酸序列及其编码蛋白BAB24651的氨基酸序列有高度同源性(＞80％)，表明NYD-SP16是一条人-小鼠同源基因。结构域分析推测NYD-SP16蛋白含有一个跨膜结构域，前期实验证实NYD-SP16蛋白是一种膜蛋白，对于睾丸发育和精子发生起着重要作用。在NYD-SP16蛋白是人-鼠高度同源的基础上，本研究采用小鼠动物模型对NYD-SP16蛋白做了进一步的功能研究。首先原核表达NYD-SP16重组蛋白，纯化后免疫小鼠制备了多克隆抗体。抗体在小鼠睾丸和精子的膜蛋白的Western Blot中均显示有特异条带，说明此抗体能识别小鼠同源蛋白。在小鼠睾丸的免疫组化中，NYD-SP16蛋白主要分布于长形精子细胞的头侧核周胞浆内。在小鼠精子其蛋白的免疫定位主要位于精子头部的顶体区域，，当精子处于不同功能状态时，CTC和免疫荧光复染显示NYD-SP16的表达在获能前后没有明显改变，但在顶体反应后明显减弱，近乎于基本消失。为检测NYD-SP16蛋白在小鼠精子获能及顶体反应中的作用，进行了在体外
[Abstract]:On the basis of differential hybridization of human testis cDNA microarray, a novel adult testis overexpression gene named NYD-SP16. was cloned in our laboratory. Sequence analysis showed that NYD-SP16 was highly homologous to the nucleotide sequence of mouse testicular gene AK006554 and the amino acid sequence of its encoding protein BAB24651 (> 80%), indicating that NYD-SP16 was a homologous gene between human and mouse. Domain analysis suggested that NYD-SP16 protein contained a transmembrane domain. Previous studies confirmed that NYD-SP16 protein is a membrane protein and plays an important role in testicular development and spermatogenesis. On the basis of the high homology of NYD-SP16 protein between human and mouse, the function of NYD-SP16 protein was further studied by using mouse animal model. Firstly, NYD-SP16 recombinant protein was expressed in prokaryotic cells, and the polyclonal antibody was prepared by immunizing mice. There were specific bands in the Western Blot of membrane protein of mouse testis and spermatozoa, which indicated that the antibody could recognize mouse homologous protein. In the immunohistochemical staining of mouse testis, NYD-SP16 protein was mainly distributed in the perinuclear cytoplasm of long sperm cells. In mouse spermatozoa, the immunological localization of its protein was mainly located in the acrosome region of the sperm head. When the sperm was in different functional states, CTC and immunofluorescence restaining showed that the expression of NYD-SP16 did not change significantly before and after capacitation, but decreased significantly after acrosome reaction. Almost disappeared. In order to detect the role of NYD-SP16 protein in mouse sperm capacitation and acrosome reaction, in vitro, the effect of NYD-SP16 protein on spermatozoa capacitation and acrosome reaction was studied.
【学位授予单位】：南京医科大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R341

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