为研究A/Anhui/1/05(H5N1)及A/Ohio/07/2009(H1N1)两株病毒神经氨酸酶(NA)茎部(stalk)区域和半胱氨酸(Cysteine,C)对流感病毒生物学特性的影响。A/Anhui/1/05(H5N1)NA(命名为AH N1)的stalk区域相对A/Ohio/07/2009(H1N1)NA存在20个氨基酸的缺失(其中包括一个半胱氨酸),将A/Ohio/07/2009(H1N1)NA(命名为09N1)的stalk区域的20个氨基酸(命名为s20)缺失构建09N1-s20,相应部分插入至H5N1的NA中构建AH N1+s20。同时为了研究stalk区域半胱氨酸对NA功能的影响,构建了AH N1+C和09N1-C(stalk区域加/减一个半胱氨酸)。运用假病毒颗粒系统研究stalk区域和半胱氨酸对病毒学特性的影响。09N1-C与09H1配对(09H1::09N1-C),其感染力比野生型09H1::09N1增强了8倍;AH N1+C与AH H5配对(AH H5::AH N1+C),其感染力比野生型AH H5::AH N1大幅度降低。09N1-s20与09H1配对(09H1::09N1-s20),其感染力为野生型09H1::09N1的4倍;AH N1+s20与AH H5配对(AH H5::AH N1+s20),感染力为野生型AH H5::AH N1的1/7。09N1-C与AH H5搭配,其感染力是AH H5::09N1的6倍;AH N1+C与09H1搭配其感染力相当低。蛋白聚合研究显示2009H1N1野生病毒的NA蛋白聚合体特征为:二聚体>>四聚体>单体;09N1-C蛋白聚合体特征为:单体为主;09N1-s20蛋白聚合体特征为:单体>>二聚体。09N1中删除半胱氨酸或敲除s20片段没有改变其蛋白的正常表达。AH N1聚合体特征以单体为主;AH N1+s20蛋白聚合特点为:二聚体>>>单体>四聚体;而AH N1+C聚合体特征为二聚体多于四聚体。研究显示:2009H1N1NA茎部区域氨基酸缺失半胱氨酸后,感染力增强;安徽H5N1的NA茎部区域添加半胱氨酸后感染力大幅降低,半胱氨酸的缺失对流感假病毒感染力增强起了重要作用。 In order to study the effects of two viral neuraminidase (NA) stalk regions and Cysteine ​​(C) pairs on A / Anhui / 1/05 (H5N1) and A / Ohio / 07/2009 (H1N1) Influenza biological effects of the virus. The stalk region of A / Anhui / 1/05 (H5N1) NA (designated AH N1) has a 20 amino acid deletion (including a cysteine) relative to A / Ohio / 07/2009 / Ohio / 07/2009 (H1N1) NA (named as 09N1) was constructed, and the corresponding part was inserted into the NA of H5N1 to construct AH N1 + s20. At the same time, AH N1 + C and 09N1-C (plus / minus one cysteine) were constructed in order to study the effect of cysteine ​​in stalk on NA function. The Effect of Stalk Region and Cysteine ​​on Virological Characteristics Using Pseudoviral Particle System. 09N1-C was paired with 09H1 (09H1 :: 09N1-C) and its infectivity was enhanced 8-fold compared to wild-type 09H1 :: 09N1; AH N1 + C was paired with AH H5 (AH H5 :: AH N1 + C) The infectivity was significantly lower than that of wild-type AH H5 :: AH N1. 09N1-s20 paired with 09H1 (09H1 :: 09N1-s20), infecting 4 times of wild type 09H1 :: 09N1; AH N1 + s20 paired with AH H5 (AH H5 :: AH N1 + s20) The wild type AH H5 :: AH N1 1 / 7.09N1-C and AH H5 with the infectivity of AH H5 :: 09N1 6 times; AH N1 + C and 09H1 with its infectivity is quite low. Protein aggregation studies showed that the 2009H1N1 wild-type virus NA protein aggregates characterized as: dimer >> tetramer> monomer; 09N1-C protein aggregates characterized by: the main monomer; 09N1-s20 protein aggregates characterized by: Monomer >> dimer. The deletion of cysteine ​​or knockout of s20 fragment in 09N1 did not change the normal expression of its protein. The characteristics of AH N1 polymer are mainly monomers; the aggregation characteristic of AH N1 + s20 protein is dimer >>> monomer> tetramer; while the AH N1 + C polymer is characterized by more dimers than tetramers . The results showed that the infectivity of 2009H1N1NA was decreased after cysteine ​​was deleted from the stem region, while the infectivity of cysteine ​​was significantly decreased after addition of cysteine ​​in the stem region of Anhui H5N1. The cysteine ​​deletion increased the virulence of influenza virus Played an important role.