牛环形泰勒虫enolase基因的原核表达及生物信息学分析

doi:10.16431/j.cnki.1671-7236.2022.10.030

Abstract

Abstract: 【Objective】 This study was aimed to obtain enolase gene from Xinjiang strain of Theileria annulata (T. annulata),and analyze the biological characteristics and reactionogenicity of the protein.【Method】 The enolase gene of T.annulata was amplified and cloned,and the prokaryotic expression vector pGEX-4T-1-enolase was constructed.The recombinant enolase protein was induced and purified.The reactivity of the recombinant enolase protein was verified by Western blotting.Bioinformatics methods were used to predict the physicochemical properties,hydrophilicity and hydrophobicity,transmembrane domain,signal peptide,phosphorylation,subcellular localization and protein interaction network of enolase gene encoded protein.【Result】 A 1 248 bp enolase gene fragment was amplified by PCR.The size of enolase recombinant protein was about 70 ku.The results of Western blotting showed that the recombinant protein reacted with the positive serum of T.annulata.enolase gene encoded 416 amino acids with a theoretical isoelectric point of 5.91 and 38 phosphorylation sites.The secondary structure was mainly composed of alpha helix (43.03%) and random coil (33.17%).It had 17 B cell epitopes,and the subcellular localization was mainly located in the cytoplasm.enolase protein interacted with phosphoglycerate mutase (PGAM),nucleoside diphosphate kinase (NDK),phosphoglycerate kinase (PGK) and heat shock protein 70 (HSP70).【Conclusion】 In this study, the enolase gene of T. annulata from Xinjiang was successfully cloned. The protein-protein interaction network predicted the protein interaction of enolase gene with glycolysis and energy metabolism. The results of this study laid a foundation for the related research on the energy metabolism pathway genes of T. annulata.

Key words: Theileria annulata; enolase gene; recombinant protein; biological characteristics

CLC Number:

S852.72

ZHENG Huizhen, PU Hao, MIAO Ronghao, GAN Lu, GE Xiaomin, Bayinchahan, LI Yongchang, GUO Qingyong. Prokaryotic Expression and Bioinformatics Analysis of Theileria annulata enolase Gene[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(10): 3972-3981.

References

[1] 张西臣,李建华.动物寄生虫病学[M].北京:科学出版社,2010. ZHANG X C,LI J H.Animal Parasitology[M].Beijing:Science Press,2010.(in Chinese)
[2] 相权珈,关贵全,刘爱红,等.环形泰勒虫SPAG蛋白的原核表达及反应原性分析[J].中国兽医科学,2020,50(3):269-275. XIANG Q J,GUAN G Q,LIU A H,et al.Prokaryotic expression and reactionogenicity analysis of recombinant Theileria annulata SPAG protein[J]. Chinese Veterinary Science,2020,50(3):269-275.(in Chinese)
[3] 海尼木古力·艾合买提,谢小婉,米热古丽·塔西铁木尔,等.新疆托克逊部分地区牛环形泰勒虫感染情况调查[J].畜牧与兽医,2017,49(6):117-119. AIHEMAITI H,XIE X W,TAXTIEMUER M,et al.Investigation on Theileria annulata infection in partial areas of Toksun,Xinjiang[J].Animal Husbandry & Veterinary Medicine,2017,49(6):117-119.(in Chinese)
[4] ZHAO S,LIU J,ZHAO H,et al.Evaluating an indirect rMPSP enzyme-linked immunosorbent assay for the detection of bovine Theileria infection in China[J].Parasitology Research,2017,116(2):667-676.
[5] 彭昱萱.烯醇化酶在微生物中的研究进展[J].科学咨询,2020,709(10):166-167. PEN Y X.Research progress of enolase in microorganisms[J].Scientific Consultation,2020,709(10):166-167.(in Chinese)
[6] PANCHOLI V.Multifunctional α-enolase:Its role in diseases[J].Cellular and Molecular Life Sciences,2001,58(7):902-920.
[7] BHOWMICK I P,KUMAR N,SHARMA S,et al.Plasmodium falciparum enolase:Stage-specific expression and sub-cellular localization[J].Malaria Journal,2009,8(1):179.
[8] GHOASH A K,MARCELO J L.Surface-expressed enolases of Plasmodium and other pathogens[J].Memorias do Instituto Oswaldo Cruz,2011,106(1):85-90.
[9] SATO K,KANO S,MATSUMOTO Y,et al.Application of yeast enolase as antigen for immunodiagnosis of malaria[J].Southeast Asian Journal of Tropical Medicine and Public Health.2000,31(1):79-84.
[10] PAL-BHOWMICK I,MEHTA M,COPPENS I,et al.Protective properties and surface localization of Plasmodium falciparum enolase[J].Infection and Immunity,2007,75(11):5500-5508.
[11] ZHANG A,BO C,MU X,et al.Identification and characterization of a novel protective antigen,enolase of Streptococcus suis serotype 2[J].Vaccine,2009,27(9):1348-1353.
[12] FENG Y J,PAN X Z,SUN W,et al.Streptococcus suis enolase functions as a protective antigen displayed on the bacterial cell surface[J].The Journal of Infectious Diseases,2009,200(10):1583-1592.
[13] BISHOP R,MUSOKE A,MORZARIA S,et al.Theileria:Intracellular protozoan parasites of wild and domestic ruminants transmitted by ixodid ticks[J].Parasitology,2004,129(7):271-283.
[14] SIODKI J,JASIK K P,KEPA M,et al.Tick-transmitted diseases caused by Apicomplexa[J].Acta Protozoologica,2011,50(2):155-161.
[15] SIDDRA A,HINES,JACOB B,et al.Exposure to ambient air causes degradation and decreased in vitro potency of buparvaquone and parvaquone[J].Veterinary Parasitology,2020,277(3):100023.
[16] NENE V,MORRISON W I.Approaches to vaccination against Theileria parva and Theileria annulata[J].Parasite Immunology,2016,38(12):724-734.
[17] DIAZ-MARTIN V,MANZANO-ROMAN R,OLEAGA A,et al.Cloning and characterization of a plasminogen-binding enolase from the saliva of the argasid tick Ornithodoros moubata[J].Veterinary Parasitology,2013,191(3-4):301-314.
[18] KIAMA T N,KIAIRA J K,KONJI V N,et al.Enzymes of glucose and glycerol catabolism in in vitro propagated Theileria parva schizonts[J].Veterinary Journal,1999,158(3):221-227.
[19] CAYIR E,ERDEMIR A,OZKAN E,et al.Cloning of intron-removed enolase gene and expression, purification,kinetic characterization of the enzyme from Theileria annulate[J].Molecular Biotechnology,2014,56(8):689-696.
[20] AUDAGNOTTO M,DAL PERARO M.Protein post-translational modifications:In silico prediction tools and molecular modeling[J].Computational and Structural Biotechnology Journal,2017,31(15):307-319.
[21] AHMED S,KABIR M,ARIF M,et al.DeepPPSite:A deep learning-based model for analysis and prediction of phosphorylation sites using efficient sequence information[J].Analytical Biochemistry,2021,612(16):113955.
[22] LEYKAUF K,TREECK M,GILSON P R,et al.Protein kinase A dependent phosphorylation of apical membrane antigen 1 plays an important role in erythrocyte invasion by the malaria parasite[J].PLoS Pathogens,2010,6(6):e1000941.
[23] PACI V,KRASTEVA I,ORSINI M,et al.Proteomic analysis of Brucella melitensis and Brucella ovis for identification of virulence factor using bioinformatics approachs[J].Molecular and Cellular Probes,2020,53(4):101581.
[24] TOMER S,CHEN K.Redundancy-weighting the PDB for detailed secondary structure prediction using deep-learning models[J].Bioinformatics,2020,36(12):3733-3738.
[25] 樊新丽,宋瑞其,呼尔查,等.牛环形泰勒虫HSP70基因的克隆表达、特性分析及互作蛋白网络构建[J].中国畜牧兽医,2021,48(1):1-11. FAN X L,SONG R Q,HU E C.Cloning,expression,characterization and protein-protein interaction analysis of Theileria annulata HSP70 gene[J].China Animal Husbandry & Veterinary Medicine,2021,48(1):1-11.(in Chinese)
[26] PALUDO G P,LORENZATTO K R,BONATTO D,et al.Systems biology approach reveals possible evolutionnarily conserved moonlighting functions for enolase[J].Computational Biology and Chemistry,2015,58(C):1-8.
[27] DIDIASOVA M,SCHAEFER L,WYGRECKA M.When place matters:Shuttling of enolase-1 across cellular compartments[J].Frontiers in Cell and Developmental Biology,2019,7(1):61.
[28] PERCONTI G,MARANTO C,ROMANCINO D P,et al.Pro-invasive stimuli and the interacting protein Hsp70 favour the route of alpha-enolase to the cell surface[J].Scientific Reports,2017,7(1):3841.

Prokaryotic Expression and Bioinformatics Analysis of Theileria annulata enolase Gene

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	WANG Leping, PAN Yan, TAN Yizhou, LI Lei, HAN Kaiou, CAO Yajie, LI Jun, ZHOU Yuqing, LIAO Yuying, MA Dongxin, GONG Yu, PENG Hao, WANG Xiaoye. Isolation,Identification and Genome Analysis of Escherichia coli Phage vB_EcoP_GN07 [J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(4): 1522-1531.
[2]	TIAN Changqing, ZHANG Kunzhong, QI Yumei, SHI Wenjing, DONG Zhijie, ZHANG Haohao, HE Zengwen, ZHI Ji, ZHAO Xuehui, CHONG Qian, XUE Huiwen, GOU Huitian. Isolation,Identification and Biological Characteristics Analysis of a Strain of Listeria monocytogenes from Environment [J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(4): 1543-1555.
[3]	CHENG Longfei, LIAO Weilian, CHEN Hongmei, JIANG Nansong, FU Qiuling, WAN Chunhe, LIU Rongchang, SHI Shaohua, HUANG Yu, FU Guanghua. Screening and Biological Characteristics of Broad-spectrum Riemerella anatipestifer Phage [J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(3): 1169-1176.
[4]	GAN Lu, ZHENG Huizhen, NUO Mingdalai, LIU Yan, JIN Min, HE Wenwen, WEN Licui, LI Yongchang, GAILIKE·Bayinchahan. Propagation of Trypanosoma evansi Yili Strain and Cloning, Expression and Bioinformatics Analysis of Its PFR Gene [J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(2): 469-478.
[5]	LIU Bin, LI Meng, XI Qianyun, SUN Jiajie, LUO Junyi, ZHANG Yongliang, CHEN Ting. Research Progress on Biological Characteristics of Hermetia illucens L. and Its Application in Poultry Production [J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(2): 586-597.
[6]	YIN Yangyan, PAN Yan, LI Xiaoning, LI Jun, LIAO Yuying, LI Changting, WEI Tianchao, PENG Hao. Study on Biological Characteristics and Antibacterial Activity in vitro of a Lactobacillus plantarum Strain [J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(2): 666-673.
[7]	GAO Yefan, SONG Hanan, WANG Yunan, WU Yue, NIU Ruili, ZONG Xianchun, GUAN Weijun. Study on the Isolation, Culture and Differentiation Potential of Limbal Stem Cells from Beijing You Chickens [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(9): 3372-3381.
[8]	ZHOU Yuqing, MA Dongxin, WU Jie, WANG Xiaoye. Isolation,Identification and Characterization Analysis of Broad-spectrum Escherichia coli O157∶H7 CVCC4050 Phage vB_EcoM_GXBP08 [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(7): 2735-2745.
[9]	YANG Zemin, LI Shuang, JIN Zhengyu, LIAO Yixiao, YANG Ying, WEN Ming. Isolation,Identification and Biological Characteristics of a Strain of Lactobacillus plantarum from Carp [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(7): 2805-2811.
[10]	LIU Jingsong, FU Jingcheng, WEN Bingyan, YANG Yanbin, GUO Shuang, JIAO Xianqin, CHEN Jin, HUANG Ruochao, WANG Yueying, LI Heping. Establishment and Analysis of Expression System of Porcine Intestinal Mucosal Protection Factor HO-1 in Bacillus pumilus [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(6): 2064-2071.
[11]	LI Jinmin, GAO Xuna, SANG Ruixin, GONG Benzhi, XU Haiyan, GU Wei, HAO Muqiang, LAN Jianghua, WANG Hong. Isolation and Evaluation of Sterilization Effect of an Avian Escherichia coli Phage [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(2): 783-190.
[12]	LI Linlin, DONG Jiawen, ZHANG Junqin, HUANG Yunzhen, SUN Minhua. Construction and Main Biological Characteristics Determination of RIA_0940 Gene Deleted Strain of Riemerella anatipestifer [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(12): 4564-4572.
[13]	LI Dapeng, CAO Chunhua, FU Pengcheng, ZHANG Jingyan, ZHANG Kang, WANG Lei, ZHANG Kai, LIU Qin, BAO Yongzhan, LI Jianxi. Identification and Biological Characteristics Evaluation of an Achromobacter Strain Isolated from Rumen of Yak [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(12): 4909-4919.
[14]	JIANG Yajun, CHEN Shiyu, XIN Ting, GUO Xiaoyu, WANG Zhaoyang, LIU Xueting, CUI Shuai, WANG Yang, ZHU Hongfei, JIA Hong. Prokaryotic Expression of African Swine Fever Virus 360-12L Protein and Preparation of Its Polyclonal Antibody [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(1): 352-360.
[15]	TANG Ning, YIN Yin, REN Huiying, ZHANG Can, LIU Wenhua. Isolation,Purification and Detection of Biological Characteristics of a High-titer and Wide-spectrum Salmonella Phage [J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(7): 2577-2583.