Mutational analysis of conserved amino acids in the fusion-promoting domain of Newcastle disease virus hemgglutinin-neuraminidase protein
10.3760/cma.j.issn.0254-5101.2012.04.016
- VernacularTitle:新城疫病毒HN糖蛋白促细胞融合区域内保守氨基酸基因突变分析
- Author:
Fulu CHU
;
Hongling WEN
;
Bin LIN
;
Chengxi SUN
;
Zhenmei LI
;
Yanyan SONG
;
Hongzhi XU
;
Zhiyu WANG
- Publication Type:Journal Article
- Keywords:
Newcastle disease virus;
Cell fusion;
Hemadeorption;
Hemagglutinin-neuraminidase
- From:
Chinese Journal of Microbiology and Immunology
2012;32(4):351-357
- CountryChina
- Language:Chinese
-
Abstract:
Objective To determine the function of conserved amino acids in the fusion-promoting domain of Newcastle disease virus (NDV) hemagglutinin-neuraminidase (HN) protein,clearly understanding mechanism of cell fusion.MethodsUsing a PCR-based site-directed mutagenesis method and the method of homology recombination occurred in vivo to change six conservative amino acids into alanine respectively.Wild type (WT) and all mutant HN proteins were exepressed in BHK-21 cells by the vacciniaT7 RNA polymerase expression system.The amount of each HN protein at the cell surface was determined by fluorescence-activated cell sorter (FACS).Cell fusion efficiency,hemadsorption activity (or receptor binding activity) and neuraminidase activity were determined.Results There was no statistic difference of cell surface expression among WT and each mutant HN protein ( P＜0.05 ).Cell fusion efficiency of each mutant protein decreased to some extent,especially 1103A decreased to 14.2％ in head.Hemadsorption activity of mutant proteins were reduced in different extent,the maximum reduction of which was also 1103A,28.2％ of wt NDV HN.There was different neuraminidase activity among each mutant HN protein.L74A increased slightly to 118.6％.L110A decreased most to 5.2％.I103A decreased second most to 5.7％.Conclusion Conserved amino acids in fusion-promoting domain of NDV HN played an important role in cell fusion.I103 was identified as a key amino acid in this domain.
