Characterizing nucleotide binding site domain (NBD) of ZzR1 resistance gene from Zingiber zerumbet: in silico ligand docking and optimizing heterologous expression

IF 1 Q3 PLANT SCIENCES

Archives of Phytopathology and Plant Protection Pub Date : 2023-08-09 DOI:10.1080/03235408.2023.2251789

A. Nair, Sumna Sasidharan

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Abstract

Abstract The Nucleotide-binding site domain (NBD) of plant resistance (R) genes plays a vital role during plant defense signaling. The functional significance of CC-NBS-LRR (Coiled coil-NBS-Leucine Rich Repeat) class of R gene designated ZzR1, characterized from Zingiber zerumbet in earlier studies, was determined by molecular modeling and docking studies. Docked complex showed the ligand GTP interacts with amino acid residues in the cleft made by GLPL and P-loop motifs of the NBD. Heterologous expression of ZzR1 NBS protein was optimized using expression vectors, pEcoli-Nterm-6xHN and pET Directional TOPO and transformed in five Escherichia coli strains namely DH5α, TOP10, BL21DE3, BL21DE3 star and BL21plysS cells. The NBS protein of 36 kDa molecular size was expressed in E. coli BL21DE3 strain using pET TOPO vector. Optimum induction was detected at 30 °C using isopropyl-1-thio-β-D-galactopyranoside (IPTG) (1 mM). The present study provides valuable information on ligand interactions and heterologous expression of ZzR1 NBD protein. graphical ABSTRACT

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泽伞形姜ZzR1抗性基因核苷酸结合位点结构域（NBD）的计算机配体对接及异源表达优化

摘要植物抗性基因的核苷酸结合位点结构域（NBD）在植物防御信号传导中起着至关重要的作用。通过分子建模和对接研究确定了CC-NBS-LRR（盘绕NBS-Leucine Rich Repeat）类R基因ZzR1的功能意义。对接复合物显示配体GTP与NBD的GLPL和P-环基序形成的裂缝中的氨基酸残基相互作用。利用表达载体pEcoli-Nterm-6xHN和pET Directional TOPO优化了ZzR1 NBS蛋白的异源表达，并将其转化到5株大肠杆菌中，即DH5α、TOP10、BL21DE3、BL21DE3星形和BL21plysS细胞。36的NBS蛋白利用pET-TOPO载体在大肠杆菌BL21DE3菌株中表达了kDa的分子量。最佳诱导时间为30 °C，使用异丙基-1-硫代-β-D-吡喃半乳糖苷（IPTG）（1 mM）。本研究为ZzR1 NBD蛋白的配体相互作用和异源表达提供了有价值的信息。图形摘要

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来源期刊

Archives of Phytopathology and Plant Protection Agricultural and Biological Sciences-Agronomy and Crop Science

CiteScore

2.20

自引率

0.00%

发文量

100

期刊介绍： Archives of Phytopathology and Plant Protection publishes original papers and reviews covering all scientific aspects of modern plant protection. Subjects include phytopathological virology, bacteriology, mycology, herbal studies and applied nematology and entomology as well as strategies and tactics of protecting crop plants and stocks of crop products against diseases. The journal provides a permanent forum for discussion of questions relating to the influence of plant protection measures on soil, water and air quality and on the fauna and flora, as well as to their interdependence in ecosystems of cultivated and neighbouring areas.