Abstract Phosphoribosylpyrophosphate synthetases 2 gene (PRPS2) is the key enzyme of biological synthesis, which plays important role in controlling tumor development, promoting tumor cell's apoptosis, inhibiting cell proliferation and promoting spermatogenesis. In order to study the role of PRPS2 gene in pig (Sus scrofa) male sterility and the function of the protein encoded by the gene, using the PRPS2 mRNA sequences of pig and related species from GenBank as reference gene sequences, Specific primers were designed and Banna mini-pig inbred line (BMI) PRPS2 gene was amplified. qRT-PCR was applied to analysis the mRNA expression profiles of 15 important tissues. Protein sequence was used to carry out functional bioinformatics analysis. cDNA sequence of 1 282 bp (GenBank accession number: KU705637, the corresponding amino acid sequence accession number: AOC89055) of BMI PRPS2 was obtained, which encodes a protein of 318 amino acids. Tissue expression profile indicated that PRPS2 gene expressed highly in the seminal vesicle gland, and low expression in the other 14 tissues. Bioinformatics analysis indicated that PRPS2 protein molecular weight 34.83 kD, and isoelectric point 6.15, contained 2 conserved domains, no transmembrane region; no signal peptide sequences, its N-terminal and the C-terminal were hydrophilic; it had 4 types of functional active sites; the probability of being located in the cytoplasm was 89%; PRPS2 gene highly conserved in evolution. The results of the study will lay a foundation for further study of the gene about its functions in pig male sterility.
[1] Ahmed M, Taylor W, Smith P R, et al. 1999. Accelerated transcription of PRPS1 in X-linked overactivity of normal human phosphoribosylpyrophosphate synthetase[J]. The Journal of Biological Chemistry, 274(11): 7482-7488. [2] Arnold K, Bordoli L, Kopp J, et al. 2006. The SWISS-MODEL workspace: a web-based environment for protein structure homology modelling[J]. Bioinformatics, 22(2): 195-201. [3] Becker M A, Heidler S A, Bell G L, et al. 1990. Cloning of cDNAs for human phosphoribosylpyrophosphate synthetases 1 and 2 and X chromosome localization of PRPS1 and PRPS2 genes[J]. Genomics, 8(3):555-561. [4] Benkert P, Biasini M, Schwede T. 2011. Toward the estimation of the absolute quality of individual protein structure models[J]. Bioinformatics, 27(3): 343-350. [5] Biasini M, Bienert S, Waterhouse A, et al. 2014. SWISS-MODEL: modelling protein tertiary and quaternary structure using evolutionary information[J]. Nucleic Acids Research, 42(Web Server issue): W252-258. [6] Cunningham J T, Moreno M V, Lodi A, et al. 2014. Protein and nucleotide biosynthesis are coupled through a single rate limiting enzyme, PRPS2, to drive cancer[J]. Cell, 157(5): 1088-1103. [7] Huo J L, Wang P, Zhao Y, et al. 2012. Molecular cloning, mRNA expression and characterization of a novel FAIM1 gene from Chinese Banna mini-pig inbred line (BMI)[J]. Journal of Animal and Veterinary Advances, 11(8): 1080-1086. [8] Ishizuka T, Iizasa T, Taira M, et al. 1992. Promoter regions of the human X-linked housekeeping genes PRPS1 and PRPS2 encoding phosphoribosylpyrophosphate synthetase subunit I and II isoforms[J]. Biochimica et Biophysica Acta, 1130(2): 139-148. [9] 鞠红. 2009. 蛋白质二级结构特征分析与相互作用预测[D]. 硕士学位论文, 哈尔滨工业大学, 导师: 王亚东. pp. 10-12.(Ju H. Protein secondary structure analysis and interaction prediction[D]. Thesis for M.S., Harbin Institute of Technology, Supervisor: Wang Y D. pp.10-12.) [10] Lee K, Keitz B, Taira M, et al. 1994. Linkage of phosphoribosylpyrophosphate synthetases 1 and 2, Prps1 and Prps2, on the mouse X chromosome[J]. Mamm Genome, 5(10): 612-615. [11] Lei B, Wan B, Peng J, et al. 2015. PRPS2 expression correlates with sertoli-cell only syndrome and inhibits the apoptosis of TM4 sertoli cells[J]. The Journal of Urology, 194(5): 1491-1497. [12] Li J, Guo W, Li F, et al. 2012. HnRNPL as a key factor in spermatogenesis: Lesson from functional proteomic studies of azoospermia patients with sertoli cell only syndrome[J]. Journal of Proteomics, 75(10): 2879-2891. [13] Li S, Lu Y, Peng B, et al. 2007. Crystal structure of human phosphoribosylpyrophosphate synthetase 1 reveals a novel allosteric site[J]. Biochemical Journal, 401(1): 39-47. [14] Mannava S, Grachtchouk V, Wheeler L J, et al. 2008. Direct role of nucleotide metabolism in C-MYC-dependent proliferation of melanoma cells[J]. Cell Cycle, 7(15): 2392-2400. [15] Nosal J M, Switzer R L, Becker M A. 1993. Overexpression, purification, and characterization of recombinant human 5-phosphoribosyl-1-pyrophosphate synthetase isozymes I and II[J]. The Journal of Biological Chemistry, 268(14): 10168-10175. [16] Taira M, Kudoh J, Minoshima S, et al. 1989. Localization of human phosphoribosylpyrophosphate synthetase subunit I and II genes (PRPS1 and PRPS2) to different regions of the X chromosome and assignment of two PRPS1-related genes to autosomes[J]. Somatic Cell and Molecular Genetics, 15(1): 29-37. [17] 王镜岩, 朱圣庚, 徐长法. 2002. 生物化学[M]. 高等教育出版社, 北京. pp. 207-220.(Wang J Y, Zhu S G, Xu C F. 2002. Biochemistry[M]. Higher Education Press, Beijing. pp. 207-220.) [18] Wang P, Huo H L, Wang S Y, et al. 2015. Cloning, sequence characterization, and expression patterns of members of the porcine TSSK family[J]. Genetics and Molecular Research, 14(4): 14908-14919. [19] 吴彬, 游锦梅, 李钰, 等. 2014. PRPS2基因shRNA质粒的构建与表达[J]. 山西医科大学学报, 45(8): 678-783. (Wu B, You J M, Li Y, et al. 2014. Construction and its expression of PRPS2 shＲNA plasmid[J]. Journal of shanxi medical university, 45(8): 678-783.) [20] 许雅鑫, 李钰, 吴彬, 等. 2015. 下调PRPS2基因表达对HCT116细胞增殖和凋亡的影响[J]. 癌变 畸变 突变, 27(2): 106-110. (Xu Y X, Li Y, Wu B, et al. 2015. Effects of down-regulated PRPS2 gene expression on HCT116 cell proliferation and apoptosis[J]. Cancerous Distortion Mutation, 27(2): 106-110.) [21] Yin Y H, Zhu X. 2003. Identification of single nucleotide polymorphisms in the promoter region of PRPS2 gene[J]. Chinese Journal of Biochemistry and Molecular Biology, 19(5): 563-565. [22] Yin Y H, Zhu X. 2001. Preliminary study of single nucleotide polymorphisms of PRPS2 gene in overproducing type of gouty patients[J]. Journal of Norman Bethune University of Medical Science, 27(3): 229-231. [23] Zeng R, Zeng Y Z. 2005. Molecular cloning and characterization of SLA-DR genes in the 133-family of the Banna mini-pig inbred line[J]. Animal Genetics, 36(3): 267-269.
