Abstract：
The glycoprotein GP64 of alphabaculovirus is crucial for viral entry and fusion. Here, we investigated the N-glycosylation patterns of Bombyx mori nucleopolyhedrovirus (BmNPV) GP64 and its signal peptide (SP) cleaved form, SPΔnGP64, along with their impacts on viral infectivity and fusogenicity. Through deglycosylation assays, we confirmed N-glycosylation of BmNPV GP64 on multiple sites. Mutational analysis targeting predicted N-glycosylation sites revealed diverse effects on viral infectivity and cell fusion. Particularly noteworthy were mutations at sites 175, which resulted in complete loss of infectivity and fusion capacity. Furthermore, LC-MS/MS analysis uncovered unexpected non-classical N-glycosylation sites, including N252, N302, N367, and N471, with only N302 and N471 identified in SPΔnGP64. Subsequent investigation highlighted the critical roles of these residues in BmNPV amplification and fusion, underscoring the essentiality of N367 glycosylation for GP64 fusogenicity. Our findings provide valuable insights into the non-classical glycosylation landscape of BmNPV GP64 and its functional significance in viral biology.
摘要：
字母杆状病毒的糖蛋白GP64对于病毒进入和融合至关重要。这里,我们研究了家蚕核型多角体病毒（BmNPV）GP64及其信号肽（SP）裂解形式的N-糖基化模式，SPΔnGP64，以及它们对病毒感染性和融合性的影响。通过去糖基化试验,我们证实了BmNPVGP64在多个位点的N-糖基化。靶向预测的N-糖基化位点的突变分析揭示了对病毒感染性和细胞融合的不同影响。特别值得注意的是位点175处的突变,其导致感染性和融合能力的完全丧失。此外，LC-MS/MS分析发现了意想不到的非经典N-糖基化位点，包括N252、N302、N367和N471,在SPΔnGP64中仅鉴定出N302和N471。随后的研究强调了这些残基在BmNPV扩增和融合中的关键作用，强调N367糖基化对GP64融合性的重要性。我们的发现为BmNPVGP64的非经典糖基化景观及其在病毒生物学中的功能意义提供了有价值的见解。
