Thaa Bastian, Kaufer Susanne, Neumann Sara A, Peibst Bernadett, Nauwynck Hans, Krause Eberhard, Veit Michael
Freie Universität Berlin, Fachbereich Veterinärmedizin, Institut für Virologie, Robert-von-Ostertag-Straße 7-13, DE-14163 Berlin, Germany.
University of Ghent, Faculty of Veterinary Medicine, Laboratory of Virology, Salisburylaan 133, BE-9820 Merelbeke, Belgium.
Virus Res. 2017 Aug 15;240:112-120. doi: 10.1016/j.virusres.2017.08.004. Epub 2017 Aug 12.
GP5 and M, the major membrane proteins of porcine reproductive and respiratory syndrome virus (PRRSV), are the driving force for virus budding and a target for antibodies. We studied co-translational processing of GP5 from an European PRRSV-1 strain. Using mass spectrometry, we show that in virus particles of a Lelystad variant, the signal peptide of GP5 was absent due to cleavage between glycine-34 and asparagine-35. This cleavage site removes an epitope for a neutralizing monoclonal antibody, but leaves intact another epitope recognized by neutralizing pig sera. Upon ectopic expression of this GP5 in cells, signal peptide cleavage was however inefficient. Complete cleavage occurred when cysteine-24 was changed to proline or an unused glycosylation site involving asparagine-35 was mutated. Insertion of proline at position 24 also caused carbohydrate attachment to asparagine-35. Glycosylation sites introduced downstream of residue 35 were used, but did not inhibit signal peptide processing. Co-expression of the M protein rescued this processing defect in GP5, suggesting a novel function of M towards GP5. We speculate that a complex interplay of the co-translational modifications of GP5 affect the N-terminal structure of the mature proteins and hence its antigenicity.
猪繁殖与呼吸综合征病毒（PRRSV）的主要膜蛋白GP5和M蛋白是病毒出芽的驱动力及抗体作用靶点。我们研究了欧洲PRRSV-1毒株中GP5的共翻译加工过程。通过质谱分析，我们发现，在莱利斯塔德病毒变异株的病毒粒子中，由于甘氨酸34和天冬酰胺35之间的切割，GP5的信号肽缺失。该切割位点去除了一种中和性单克隆抗体的表位，但另一个可被猪中和血清识别的表位保持完整。然而，当该GP5在细胞中异位表达时，信号肽切割效率低下。当半胱氨酸24变为脯氨酸或涉及天冬酰胺35的一个未使用的糖基化位点发生突变时，会发生完全切割。在第24位插入脯氨酸也会导致天冬酰胺35发生碳水化合物连接。在第35位残基下游引入的糖基化位点可被利用，但不抑制信号肽加工。M蛋白的共表达挽救了GP5的这种加工缺陷，提示M蛋白对GP5具有新功能。我们推测，GP5共翻译修饰的复杂相互作用影响成熟蛋白的N端结构，进而影响其抗原性。
