极端温度和pH条件下乙型肝炎表面蛋白的抗原和物理化学表征

StephenW

Vaccine. 2019 Sep 10. pii: S0264-410X(19)31193-4. doi: 10.1016/j.vaccine.2019.09.005. [Epub ahead of print]
Antigenic and physicochemical characterization of Hepatitis B surface protein under extreme temperature and pH conditions.
Lopes JLS1, Oliveira DCA2, Utescher CLA2, Quintilio W2, Tenório ECN2, Oliveira CLP3, Fantini MCA3, Rasmussen MK4, Bordallo HN5, Sant'Anna OA2, Botosso VF2.
Author information

1
    Institute of Physics, University of São Paulo, São Paulo, Brazil. Electronic address: [email protected].
2
    Butantan Institute, São Paulo, Brazil.
3
    Institute of Physics, University of São Paulo, São Paulo, Brazil.
4
    Niels Bohr Institute, University of Copenhagen, Copenhagen, Denmark; Department of Health Technology, Technical University of Denmark, Lyngby, Denmark.
5
    Niels Bohr Institute, University of Copenhagen, Copenhagen, Denmark; European Spallation Source, Lund, Sweden.

Abstract

Hepatitis B virus causes acute and chronic infections in millions of people worldwide and, since 1982, a vaccine with 95% effectiveness has been available for immunization. The main component of the recombinant hepatitis B vaccine is the surface antigen protein (HBsAg). In this work, the effect of pH, ionic strength and temperature on the native state of the HBsAg antigen were studied by a combination of biophysical methods that included small angle X-ray scattering, synchrotron radiation circular dichroism, fluorescence and surface plasmon resonance spectroscopies, as well as in vivo and in vitro potency assays. The native conformation, morphology, radius of gyration, and antigenic properties of the HBsAg antigen demonstrate high stability to pH treatment, especially in the pH range employed in all stages of HBsAg vaccine production and storage. The HBsAg protein presents thermal melting point close to 56 °C, reaching a more unfolded state after crossing this point, but it only experiences loss of vaccine potency and antigenic properties at 100 °C. Interestingly, a 6-month storage period does not affect vaccine stability, and the results are similar when the protein is kept under refrigerated conditions or at room temperature (20 °C). At frozen temperatures, large aggregates (>200 nm) are formed and possibly cause loss of HBsAg content, but that does not affect the in vivo assay. Furthermore, HBsAg has a well-ordered secondary structure content that is not affected when the protein is formulated with silica SBA-15, targeting the oral delivery of the vaccine. The combined results from all the characterization techniques employed in this study showed the high stability of the antigen at different storage temperature and extreme values of pH. These findings are important for considering the delivery of HBsAg to the immune system via an oral vaccine.

Copyright © 2019 Elsevier Ltd. All rights reserved.
KEYWORDS:

HBsAg antigenicity; Hepatitis B; Thermal stability; pH stability

PMID:
    31519445
DOI:
    10.1016/j.vaccine.2019.09.005

StephenW

疫苗。 2019年9月10日.pii：S0264-410X（19）31193-4。 doi：10.1016 / j.vaccine.2019.09.005。 [印刷前的电子版]
极端温度和pH条件下乙型肝炎表面蛋白的抗原和物理化学表征。
Lopes JLS1，Oliveira DCA2，Utescher CLA2，Quintilio W2，TenórioECN2，Oliveira CLP3，Fantini MCA3，Rasmussen MK4，Bordallo HN5，Sant'Anna OA2，Botosso VF2。
作者信息

1
巴西圣保罗圣保罗大学物理研究所。电子地址：[email protected]。
2
巴西圣保罗Butantan研究所。
3
巴西圣保罗圣保罗大学物理研究所。
4
丹麦哥本哈根大学Niels Bohr研究所;丹麦技术大学卫生技术系，丹麦Lyngby。
五
丹麦哥本哈根大学Niels Bohr研究所;欧洲散裂源，瑞典隆德。

抽象

乙型肝炎病毒引起全世界数百万人的急性和慢性感染，自1982年以来，已有一种效率为95％的疫苗可用于免疫接种。重组乙型肝炎疫苗的主要成分是表面抗原蛋白（HBsAg）。通过结合生物物理方法研究了pH，离子强度和温度对HBsAg抗原天然状态的影响，包括小角度X射线散射，同步辐射圆二色性，荧光和表面等离子共振光谱等。体内和体外效力测定。 HBsAg抗原的天然构象，形态，回转半径和抗原性质证明了对pH处理的高稳定性，尤其是在HBsAg疫苗生产和储存的所有阶段中使用的pH范围内。 HBsAg蛋白的热熔点接近56°C，在穿过这一点后达到更加展开的状态，但只有exp有趣的是，6个月的储存期不会影响疫苗的稳定性，并且当保留蛋白质时结果相似在冷藏条件下或室温下（20°C）。在冷冻温度下，形成大的聚集体（> 200nm）并且可能导致HBsAg含量的损失，但这不会影响体内测定。此外，HBsAg具有良好有序的二级结构含量，当蛋白质用二氧化硅SBA-15配制时，其目标是口服递送疫苗。本研究中使用的所有表征技术的组合结果显示了抗原在不同储存温度和极端pH值下的高稳定性。通过口服疫苗将HBsAg递送至免疫系统。

版权所有©2019 Elsevier Ltd.保留所有权利。
关键词：

HBsAg抗原性;乙型肝炎;热稳定性; pH稳定性

结论：
31519445
DOI：
10.1016 / j.vaccine.2019.09.005