噬菌体Bp7蛋白gp38的克隆、表达及其受体识别活性分析

doi:10.11843/j.issn.0366-6964.2018.11.018

Abstract

Abstract:

To identify the role of protein gp38 in the adsorption process of phage Bp7, gp38 gene was amplified by PCR, and the amino acid sequence was characterized. The recombinant plasmid pColdTF-gp38 was constructed and expressed in E. coli BL21, then identified by SDS-PAGE and Western blot. Protein gp38 was purified by affinity chromatography to prepare polyclonal antibody. Immunoelectron microscopy assay was used to locate gp38 on phage Bp7, protein competition assay and antibody blocking assay were used to determine gp38 and its antibody effects on phage Bp7 adsorption efficiency. Amino acid sequence analysis of phage Bp7 protein gp38 showed that there was no homology with T4 bacteriophage, but shared homology with phage T2 and T6. The conserved regions and variable regions of gp38 C-terminal sequence were arranged at intervals, showing a mosaic characteristics; the recombined plasmid pColdTF-gp38 was constructed and expressed in E. coli BL21, The gp38 polyclonal antibody was prepared with a 1:16 titer. The immunoelectron microscopy showed that the gp38 antibody could bind to the long tail fiber and cause the aggregation of phage Bp7; protein competition test and antibody blocking test showed that gp38 and its antibody could completely inhibit the absorption of phage Bp7 to host strain E. coli K12. All of results indicated that gp38 is the tail fiber protein of phage Bp7, located at the end of the long tail fiber, and played a key role as the receptor recognition protein in phage Bp7 adsorption process, which further provided the theoretical foundation to elucidate the receptor recognition mechanism of phage Bp7.

Key words: bacteriophage Bp7, gp38 protein, sequence analysis, expression, immunoelectron microscopy, receptor-recognizing activity

CLC Number:

Q939.48

MA Hai-lan, SUN Hu-zhi, REN Hui-ying, ZHANG Can. Cloning, Expression and Receptor-Recognizing Activity of Bacteriophage Bp7 Protein gp38[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(11): 2461-2467.

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Cloning, Expression and Receptor-Recognizing Activity of Bacteriophage Bp7 Protein gp38

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