Russell Bonnie Leigh, Gildenhuys Samantha
Department of Life and Consumer Sciences, College of Agriculture and Environmental Sciences, University of South Africa, Florida, South Africa.
Clin Exp Vaccine Res. 2020 Jul;9(2):108-118. doi: 10.7774/cevr.2020.9.2.108. Epub 2020 Jul 31.
The Bluetongue virus (BTV) is an economically significant disease that affects mainly wild and domestic ruminants. BTV is most often seen symptomatically in sheep, but is easily carried by goats, cattle, and wild ruminants. To date there are several problems with the vaccines currently available for BTV, and one of the most promising candidates to increase vaccine efficacy is a protein-based vaccine, for which viral protein 7 (VP7) is a great candidate to be included in it. In order to further these studies, the stability of BTV VP7 in common vaccine additives needs to be investigated.
Recombinant BTV VP7 was expressed in a bacterial cell system and purified before being analysed using spectroscopic techniques including far-ultraviolet (UV) circular dichroism and intrinsic tryptophan fluorescence. BTV was analysed in a number of different buffer conditions.
We report here that BTV VP7 maintains its native secondary structure until at least 52℃ and native-like tertiary structure to at least 80℃. Far-UV circular dichroism and intrinsic tryptophan fluorescence emission spectra indicate significant secondary and tertiary structure remaining even at 90℃, respectively. Six M guanidinium chloride is able to unfold BTV VP7 while 8 M urea could not.
Twenty percent glycerol and 300 mM sodium chloride appear to have a protective effect on BTV VP7's structure, as significantly more structure is seen at 90℃ when compared to BTV VP7 without the addition of these chemicals. Both glycerol and sodium chloride are common vaccine additives.
蓝舌病毒（BTV）是一种具有重要经济影响的疾病，主要感染野生和家养反刍动物。BTV在绵羊中最常表现出症状，但山羊、牛和野生反刍动物很容易携带该病毒。迄今为止，目前可用的BTV疫苗存在几个问题，提高疫苗效力最有前景的候选方法之一是基于蛋白质的疫苗，病毒蛋白7（VP7）是其中非常适合包含的一种候选蛋白。为了推进这些研究，需要研究BTV VP7在常见疫苗添加剂中的稳定性。
重组BTV VP7在细菌细胞系统中表达并纯化，然后使用包括远紫外（UV）圆二色性和色氨酸固有荧光在内的光谱技术进行分析。在多种不同缓冲条件下对BTV进行分析。
我们在此报告，BTV VP7至少在52℃时保持其天然二级结构，至少在80℃时保持类似天然的三级结构。远紫外圆二色性和色氨酸固有荧光发射光谱分别表明，即使在90℃时仍有显著的二级和三级结构保留。6M盐酸胍能够使BTV VP7解折叠，而8M尿素则不能。
20%甘油和300mM氯化钠似乎对BTV VP7的结构有保护作用，因为与未添加这些化学物质的BTV VP7相比，在90℃时能看到更多的结构。甘油和氯化钠都是常见的疫苗添加剂。
