Human ribonuclease 9, a member of ribonuclease A superfamily, specifically expressed in epididymis, is a novel sperm-binding protein
Human ribonuclease 9, a member of ribonuclease A superfamily, specifically expressed in epididymis, is a novel sperm-binding protein
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DOI：
10.1038/aja.2008.30
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发表时间：
2009-03-01
影响因子：
2.9
通讯作者：
Shi, Guang-Xia
中科院分区：
文献类型：
--
作者：
Cheng, Gui-Zhi;Li, Jian-Yuan;Shi, Guang-Xia
为了探究人核糖核酸酶9 （RNase9）的功能，我们构建了哺乳动物融合表达载体pcDNA-hRNase9，从HEK293T细胞中制备了重组人RNase9 - his融合蛋白，并测定了其n端氨基酸序列。根据测定的成熟蛋白，在大肠杆菌中制备了重组人RNase 9。检测重组人RNase 9的溶核糖活性和抑菌活性，并采用间接免疫荧光法分析重组人RNase 9在组织、射精精子和体外获能精子上的分布。结果表明，重组人RNase 9对酵母tRNA没有可检测到的核糖核溶解活性，但对大肠杆菌有一定的抑菌活性，且呈浓度/时间依赖性。免疫荧光分析显示，预测的人类RNase 9存在于整个附睾，但不存在于所检查的其他组织中，人类RNase 9也存在于人类射精精子和体外获能精子的整个头部和颈部区域。这些结果提示人类RNase 9可能在男性生殖道宿主防御中发挥作用。
To explore the functions of human ribonuclease 9 ( RNase 9), we constructed a mammalian fusion expression vector pcDNA-hRNase9, prepared recombinant human RNase 9-His fusion protein from HEK293T cells and determined its N-terminal amino acid sequences. According to the determined mature protein, recombinant human RNase 9 was prepared in E. coli. Ribonucleolytic activity and antibacterial activity of recombinant human RNase 9 were detected, and the distribution of human RNase 9 on tissues and ejaculated spermatozoa and in vitro capacitated spermatozoa were analyzed via indirect immunofluorescence assay. The results showed that recombinant human RNase 9 did not exhibit detectable ribonucleolytic activity against yeast tRNA, but exhibited antibacterial activity, in a concentration/time dependent manner, against E. coli. Immunofluorescent analyses showed that the predicted human RNase 9 was present throughout the epididymis, but not present in other tissues examined, and human RNase 9 was also present on the entire head and neck regions of human ejaculated spermatozoa and in vitro capacitated spermatozoa. These results suggest that human RNase 9 may play roles in host defense of male reproductive tract.