猫疱疹病毒1型病毒样颗粒在酵母中的制备及体外组装OA北大核心CSTPCD
Preparation of feline herpesvirus type 1 VLPs in yeast and assembled in vitro
为研发猫疱疹病毒1型(FHV-1)病毒样颗粒(VLPs)疫苗,将FHV-1 gD基因和自组装蛋白1301基因序列融合后克隆到pPink-HC载体中,由毕赤酵母表达重组gD蛋白.随后观察不同pH对蛋白质表达和VLPs组装效果的影响.结果显示,经SDS-PAGE和Western-blot鉴定,发现培养基pH3.0条件下毕赤酵母表达出了完整的重组gD蛋白,而pH6.0条件下表达的蛋白分子质量小于预计大小;另外,组装液pH8.0时VLPs的组装效果优于pH6.0、7.0.结果表明,酸性环境可有效抑制毕赤酵母内蛋白酶对重组gD蛋白的水解作用,重组gD蛋白可在弱碱性环境中利用自组装蛋白1301在体外完成组装,为FHV-1 VLPs疫苗制备奠定了基础.
To develop virus-like particles(VLPs)vaccines against feline herpesvirus type 1(FHV-1),FHV-1 gD gene was fused with self-assembling protein 1301 gene and then cloned into pPink-HC vector.The recombinant gD protein was expressed in Pichia pastoris.The effects of different pH on protein expres-sion and VLPs assembly were then observed.The results showed that P.pastoris expressed intact recombi-nant gD protein at pH3.0,which was identified by SDS-PAGE and Western-blot,while the molecular weight of protein expressed under pH6.0 condition was smaller than expected.In addition,the assembly effect of VLPs at pH8.0 was better than that of pH6.0 and 7.0.These results showed that acidic environment could effectively inhibit the hydrolysis of recombinant gD protein by proteases in P.pastoris.Recom-binant gD protein can be assembled into VLPs in vi tro by using self-assembled protein 1301 in a weak al-kaline environment,which lays a foundation for preparation of FHV-1 VLPs vaccine.
赵梦杉;吕熙;王运航;裴辰辰;刘艺佳;郭慧琛;孙世琪;杨丰利;董虎
长江大学动物科学技术学院,湖北荆州 434025||中国农业科学院兰州兽医研究所兰州大学动物医学与生物安全学院动物疫病防控全国重点实验室,甘肃兰州 730000中国农业科学院兰州兽医研究所兰州大学动物医学与生物安全学院动物疫病防控全国重点实验室,甘肃兰州 730000||榆林学院生命科学学院,陕西榆林 719000中国农业科学院兰州兽医研究所兰州大学动物医学与生物安全学院动物疫病防控全国重点实验室,甘肃兰州 730000长江大学动物科学技术学院,湖北荆州 434025
畜牧业
猫疱疹病毒1型病毒样颗粒酵母表达系统gD基因
feline herpesvirus type 1virus-like particlesyeast expression systemgD gene
《中国兽医科学》 2024 (007)
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国家重点研发计划项目(2022YFD1800603,2021YFD1800300);国家自然科学基金项目(32072859,32072847,32002272)
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