BACKGROUND The paramyxovirus haemagglutinin-neuraminidase (HN) is a multifunctional protein that is responsible for attachment to receptors, removal of receptors from infected cells to prevent viral self-aggregation (neuraminidase, NA) and fusion promotion. It is commonly accepted that there are two receptor binding sites in the globular head of HN, and the second receptor binding site is only involved in the function of receptor binding and fusion promotion. METHODS 10 conserved residues in the second receptor binding site of Newcastle disease virus (NDV) HN were chosen and substituted to alanine (A). The desired mutants were examined to detect the functional change in hemadsorption (HAD) ability, NA activity and fusion promotion ability. RESULTS The HAD and fusion promotion ability of mutants C172A, R174A, C196A, D198A, Y526A and E547A were abolished. Compared with wild-type (wt) HN, the HAD of mutants T167A, S202A and R516A decreased to 55.81, 44.53, 69.02%, respectively, and the fusion promotion ability of these three mutants decreased to 54.74, 49.46, 65.26%, respectively; however, mutant G171A still maintained fusion promotion ability comparable with wt HN but had impaired HAD ability. All the site-directed mutations altered the NA activity of NDV HN without affecting protein cell surface expression. CONCLUSIONS The data suggest that mutants C172A, R174A, C196A, D198A, Y526A and E547A do not allow the conformational change that is required for fusion promotion ability and HAD activity, while the other mutants only affect the conformational change to a limited extent, except mutant G171A with intact fusion promotion ability. Overall, the conserved amino acids in the second receptor binding site, especially residues C172, R174, C196, D198, Y526 and E547, are crucial to normal NDV HN protein function.

中文翻译：

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高度保守的氨基酸在新城疫病毒HN蛋白的第二个受体结合位点中的作用。

背景技术副粘病毒血凝素神经氨酸酶（HN）是一种多功能蛋白，其负责与受体的连接，从感染细胞中去除受体以防止病毒自身聚集（神经氨酸酶，NA）和融合促进。普遍认为，HN的球状头部有两个受体结合位点，而第二个受体结合位点仅参与受体结合和融合促进的功能。方法选择新城疫病毒（NDV）HN第二受体结合位点中的10个保守残基，并将其替换为丙氨酸（A）。检查所需的突变体以检测溶血（HAD）能力，NA活性和融合促进能力的功能变化。结果突变体C172A，R174A，C196A，D198A的HAD和融合促进能力 取消了Y526A和E547A。与野生型（wt）HN相比，突变体T167A，S202A和R516A的HAD分别降低至55.81、44.53、69.02％，这三个突变体的融合促进能力分别降低至54.74、49.46、65.26％。然而，突变体G171A仍然保持与wt HN相当的融合促进能力，但是损害了HAD能力。所有定点突变都改变了NDV HN的NA活性，而不会影响蛋白质细胞表面的表达。结论数据表明突变体C172A，R174A，C196A，D198A，Y526A和E547A不允许融合促进能力和HAD活性所需的构象变化，而其他突变体仅在有限的程度上影响构象变化，除了突变体具有完整融合促进能力的G171A。全面的，