Klink Holly A, Brady Ryan P, Topliff Christina L, Eskridge Kent M, Srikumaran Subramaniam, Kelling Clayton L
Department of Veterinary and Biomedical Sciences, University of Nebraska, East Campus Loop and Fair Street, Lincoln, NE 68583-0905, USA.
Vaccine. 2006 Apr 12;24(16):3388-95. doi: 10.1016/j.vaccine.2005.12.067. Epub 2006 Feb 6.
Bovine respiratory syncytial virus (BRSV) is an etiological component of the bovine respiratory tract disease complex. Infection with BRSV following vaccination, or re-infection following natural infection is common since protection is incomplete. The objectives of this study were to create plasmid DNA constructs encoding single or multiple N-glycosylation-site deletion BRSV fusion (F) proteins, and evaluate their expression in cell culture, and potential to induce anti-BRSV F antibody responses in BALB/c mice. Four plasmid DNAs were constructed, each encoding 1-4 N-glycosylation-site deletions: Gly4, Gly2/4, Gly1/2/4 and Gly1/2/3/4. Each of the N-glycosylation-site deletion BRSV F proteins were expressed in COS-7 cells following transfection with plasmid DNA. Inoculation of BALB/c mice with plasmid DNA, resulted in a significant anti-BRSV F IgG response to the wild-type (WT) F and glycosylation-site deletion protein Gly2/4. Gly2/4 elicited a higher antibody titer than the fully glycosylated WT F protein. Significant neutralizing antibody titers were detected following immunization with the Gly2/4 plasmid DNA. These glycosylation-site deletion BRSV F proteins will be useful to characterize the effects of glycosylation on immunogenicity in the natural host, and may lead to a new approach for the generation of BRSV vaccines.
牛呼吸道合胞病毒（BRSV）是牛呼吸道疾病综合征的一个病因成分。接种疫苗后感染BRSV，或自然感染后再次感染很常见，因为保护并不完全。本研究的目的是构建编码单个或多个N-糖基化位点缺失的BRSV融合（F）蛋白的质粒DNA构建体，评估它们在细胞培养中的表达，以及在BALB/c小鼠中诱导抗BRSV F抗体反应的潜力。构建了四种质粒DNA，每种编码1-4个N-糖基化位点缺失：Gly4、Gly2/4、Gly1/2/4和Gly1/2/3/4。用质粒DNA转染后，每个N-糖基化位点缺失的BRSV F蛋白都在COS-7细胞中表达。用质粒DNA接种BALB/c小鼠，导致对野生型（WT）F和糖基化位点缺失蛋白Gly2/4产生显著的抗BRSV F IgG反应。Gly2/4诱导的抗体滴度高于完全糖基化的WT F蛋白。用Gly2/4质粒DNA免疫后检测到显著的中和抗体滴度。这些糖基化位点缺失的BRSV F蛋白将有助于表征糖基化对天然宿主免疫原性的影响，并可能导致一种产生BRSV疫苗的新方法。
