版纳微型猪近交系PPP2CA克隆、表达及蛋白质功能生物信息学分析

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摘要蛋白磷酸酶2A(protein phosphatases of the type 2A, PP2A)是一种广泛表达的丝氨酸/苏氨酸磷酸酶，介导蛋白质的去磷酸化从而调控多种关键信号通路的活性。该酶由催化亚基、结构亚基、调节亚基形成异源三聚体复合物发挥作用。本研究借助GenBank中猪(Sus scrofa)及其他物种的蛋白磷酸酶2A催化亚基α (protein phosphatase 2A catalytic subunit α, PPP2CA) mRNA序列，设计特异引物扩增版纳微型猪近交系(Banna mini-pig inbred line, BMI)PPP2CA基因编码区序列，并对其进行表达和功能特性分析。应用qRT-PCR分析组织mRNA表达谱，并对编码的蛋白质序列进行生物信息学分析。扩增得到了BMI PPP2CA 930 bp (GenBank No. KU705627)的完整CDS序列，共编码309个氨基酸。与其他组织相比PPP2CA基因在尿道球腺和精囊腺中极显著高表达(P＜0.01)，在睾丸中表达量也相对较高，在肝、结肠、脾、肺、十二指肠、前列腺、肾、附睾及脑中中度表达；在胃、心、肌肉中表达水平极显著低于其他组织。PPP2CA蛋白质功能预测表明其存在1个保守结构域MPP_PP2A_PP4_PP6，存在4类功能活性位点，无跨膜螺旋结构，无信号肽序列，N末端和C末端均亲水，位于细胞质的概率是94.1%。二级结构预测表明该蛋白α-螺旋含量最高，是组成N端的主要结构，C端有一段无规则卷曲，多物种氨基酸序列比对结果显示，PPP2CA序列在不同物种间相似度很高，推测其在进化中高度保守，为深入研究PPP2CA基因在猪精子获能方面作用机制提供了资料基础。

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	王淑燕
	张霞
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	霍海龙
	李卫真

关键词 ：蛋白磷酸酶2A催化亚基A, 版纳微型猪近交系, 雄性育性, 生物信息学

Abstract：Protein phosphatase 2A (PP2A) is a ubiquitously expressed serine-threonine phosphatase mediating protein dephosphorylation to regulate the activities of many key pathways. PP2A is a heterotrimeric including catalytic subunit, structural subunit and regulatory subunit. To clone the coding sequence and analyze the character of expression and function of protein phosphatase 2A catalytic subunit α gene (PPP2CA), the specific primers of PPP2CA gene of Banna mini-pig (Sus scrofa) inbred line (BMI) was designed and the coding sequence was amplified using the PPP2CA mRNA sequences of pig and other species from GenBank as reference sequences. qRT-PCR was used to detect expression profiles of tissues mRNA. At the same time, the amino acid sequences were analysed by functional bioinformatics. A complete coding sequence of 930 bp of BMI PPP2CA was obtained, which encoded 309 amino acids. Comparing with other tissues, the PPP2CA gene expression in the urethral ball glands and seminal vesicle appeared extremely significance (P＜0.01). The expression in the testis was higher than other tissues. It was expressed moderately in the liver, colon, spleen, lung, duodenum, prostate, kidney, epididymis and brain, weakly in the stomach, heart and muscle. Functional prediction of PPP2CA protein indicated that the PPP2CA contained one conserved domain MPP_PP2A_PP4_PP6 and four kinds functional active sites, without transmembrane helix and signal peptide. The N-terminal and C-terminal were hydrophilic and located in cytoplasmic with 94.1%. It was predicted that the α helices content of the second structure were the highest which were the major structure of the N terminal, while the random coils were the major structure of the C terminal. The high similarity of PPP2CA amino acids alignment between BMI and other species indicated that this gene was highly conservative in evolution. This research can lay a further foundation for clarifying mechanism of the PPP2CA gene on sperm capacitation.

Key words： PP2CA Banna mini-pig inbred line (BMI) Male fertility Bioinformatics

收稿日期: 2018-02-20 出版日期: 2018-09-26

ZTFLH:

S 828. 2

基金资助:版纳微型猪近交系生精障碍关键基因pSeptin12、14的功能与作用机制研究

通讯作者: 霍金龙 E-mail: jinlonghuo973@163.com

引用本文:

王淑燕张霞霍金龙王配霍海龙李卫真. 版纳微型猪近交系PPP2CA克隆、表达及蛋白质功能生物信息学分析[J]. , 2018, 26(10): 1737-1746.

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http://journal05.magtech.org.cn/Jwk_ny/CN/ 或 http://journal05.magtech.org.cn/Jwk_ny/CN/Y2018/V26/I10/1737

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