Affinity Maturation of Anti-carbofuran Single Chain Fv by Site-directed Mutagenesis

doi:10.7503/cjcu20120308

Abstract

Abstract:

To obtain higher affinity anti-carbofuran(CBF) single-chainfragment(scFv) antibody, the scFv model by homology modeling was constructed according to the amino acids sequence and the active pocket region were determined. Through molecular docking model of CBF and scFv, it was found that hydrophobic interaction and hydrogen bonding played an important role of antibody affinity. Then the hydrophilic amino acids HArg40 and LHis38 were changed to hydrophobic amino acids. The molecular docking of CBF and mutant antibody indicated that higher docking scores could be obtained when the leucine was taken as the mutation amino acid. Furthermore, mutation scFv gene was constructed, soluble recombinant protein was expressed and the affinity of evolved scFv(evoscFv) was identified by ELISA. The results showed that the 50% inhibition of binding of evoscFv against CBF was 18.11 μg/L which was lower than the wild type scFv antibody, and the affinity was also improved 2.23-fold. All these indicated that directed affinity maturation anti-CBF evoscFv was obtained through the analysis of molecule docking and replacement of target amino acid residues.

Key words: Carbofuran, Single-chain antibody, Molecular docking, Site-directed mutagenesis, Affinity maturation

CLC Number:

O629

TrendMD:

DENG Long, XIAO Zhi-Li, DONG Jie-Xian, WANG Hong, YANG Wu-Ying, XIE Xi, YANG Jin-Yi, SHEN Yu-Dong, SUN Yuan-Ming. Affinity Maturation of Anti-carbofuran Single Chain Fv by Site-directed Mutagenesis[J]. Chem. J. Chinese Universities, 2012, 33(11): 2486.

[1]	LEI Xiaotong, JIN Yiqing, MENG Xuanyu. Prediction of the Binding Site of PIP₂ in the TREK-1 Channel Based on Molecular Modeling [J]. Chem. J. Chinese Universities, 2021, 42(8): 2550.
[2]	SHUAI Die, ZHAO Meijuan, CHEN Bingnian, WANG Li. Inhibitory Effect of Four Kinds of Keegin-type Phosphomolybdate on Tyrosinase and Melanin Formation and Its Antioxidant Activities [J]. Chem. J. Chinese Universities, 2021, 42(12): 3579.
[3]	YANG Ju, SU Lijiao, LI Canhua, LU Jiajia, YANG Junli, GU Jie, YANG Li, YANG Lijuan. Host-guest Complexation Behavior of Nardosinone and Water-soluble Phosphate Salt Pillar［6］arene [J]. Chem. J. Chinese Universities, 2021, 42(10): 3099.
[4]	ZHANG Aiqin, WANG Man, SHEN Gangyi, JIN Jun. Interactions Between Polybrominated Diphenyl Ethers and Human Serum Albumin Using SPR and Molecular Docking [J]. Chem. J. Chinese Universities, 2020, 41(9): 2054.
[5]	WANG Lianping,LI Qingjie,LIU Xiaoyan,REN Yueying,YANG Xiuwei. Screening of Cholinesterase Inhibitors in Fructus Evodiae Alkaloids Based on UFLC-MS/molecular Simulation ^† [J]. Chem. J. Chinese Universities, 2020, 41(1): 111.
[6]	WANG Xiaoxia, MA Litong, NIE Zhihua, WANG Zhengde, CUI Jinlong, ZHAO Wenyuan, SAI Huazheng. Interaction Between Fulvic Acid and Pepsin Investigated by Multispectral and Molecular Docking Simulation ^† [J]. Chem. J. Chinese Universities, 2019, 40(9): 1840.
[7]	LIU Zhongcheng, LIU Shifang, ZHANG Su, YANG Yanlei, LI Fei, ZHANG Nan, YUAN Xin, ZHANG Yanfen. Structure Prediction and Screening of Oligonucleotide Aptamers Target Cε3-Cε4 Protein^† [J]. Chem. J. Chinese Universities, 2019, 40(1): 83.
[8]	XIN Meiling, CHU Zhenhua, LI Yu. Molecular Modification of Polychlorinated Biphenyl Dihydroxy Derivatives Through Molecular Docking Associated with CoMSIA/HQSAR Models^† [J]. Chem. J. Chinese Universities, 2018, 39(2): 299.
[9]	WANG Yan, CHEN Ping, WANG Yunfei, LIU Guiying, YANG Xi, SU Ying, LI Junyang, LIU Weiwei, LIN Lie. Spectral Characterization of the Interaction Between Methamphetamine and Serum Albumin^† [J]. Chem. J. Chinese Universities, 2018, 39(11): 2507.
[10]	DUAN Yongbin, YIN Yan, MENG Fanli, ZHAO Lianhua, LIU Yukun, YUAN Zhe, FENG Yangbo. Design, Synthesis and Biological Evaluation of Benzothiazoles as Highly Potent ROCK Inhibitors Through Molecular Docking and Free Energy Calculations^† [J]. Chem. J. Chinese Universities, 2017, 38(9): 1568.
[11]	WANG Song, GUAN Shanshan, WAN Yongfeng, SHAN Yaming, ZHANG Hao. Molecular Dynamics Simulation Study on the Binding Modes of Angiotensin-converting Enzyme with Inhibitory Peptides^† [J]. Chem. J. Chinese Universities, 2017, 38(7): 1216.
[12]	TANG Qian, SU Jinhong, CAO Hongyu, WANG Lihao, SHI Fei, WANG Ailing, GONG Tingting, JIN Xiaojun, ZHENG Xuefang. Interaction of Pyrimidine Derivatives with Human Serum Albumin^† [J]. Chem. J. Chinese Universities, 2017, 38(11): 1982.
[13]	ZHAO Bin, HUO Jingqian, XING Jihong, QI Meng, ZHANG Jinlin, DONG Jingao. Homologous Modeling of Transketolase AtTKL1 and Its Combination with α-Terthienyl in Arabidopsis Thaliana^† [J]. Chem. J. Chinese Universities, 2015, 36(4): 682.
[14]	DONG Lu, YI Zhongsheng, WU Zhiwei, WANG Haiyang, ZHANG Aiqian. Mechanism Study on the Interaction Between 2'-Hydroxy-2,4-dibromo Diphenyl Ethers and Human Serum Albumin Based on Spectroscopic Methods and Computional Simulations^† [J]. Chem. J. Chinese Universities, 2015, 36(3): 516.
[15]	ZHANG Min, ZHANG Yi, LI Chengtao, QIN Jiaxiang, Gao Rang, MA Xiaoning, QIU Jianhui. N435 Enzymatic Degradation Difference and Molecular Modeling of Hydrophilic Modified PBS-based Copolymer^† [J]. Chem. J. Chinese Universities, 2015, 36(3): 568.

Affinity Maturation of Anti-carbofuran Single Chain Fv by Site-directed Mutagenesis

PDF (PC)

Knowledge

Cited

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments