Molecular Assembly and Stability of Human Hepatitis B Virus Surface Antigen Vaccine

现在原则上可以利用基因工程技术生产优于常规疫苗的上级疫苗。然而，从酵母中获得的人B型肝炎病毒表面抗原疫苗（γ-HBsAg疫苗）是将工程技术引入疫苗生产的第一个也是唯一一个成功的结果。其功效被认为是源自主要由蛋白质多肽和脂质组成的组装体的特定性质和高稳定性。本研究项目的动机来自我们的信念，即必须了解组分的组装模式和稳定性的来源，以提供基础知识，希望通过该方法生产其他有效的疫苗。发现γ-HBsAg疫苗颗粒是球形的，并且根据动态光散射判断，其半径约为19 nm，小角X射线散射和电子显微镜。通过小角X射线散射在疫苗颗粒中未检测到规则结构。疫苗颗粒从TSK-Gel G600 OPW柱上洗脱为对称峰。对洗脱的监测给出了本研究中目标疫苗颗粒的摩尔质量估计值为5，040 kg/mol，与病毒表面抗原的多肽相比，其组成多肽在N-末端侧具有9个额外的氨基酸残基。由后一种多肽组成的疫苗颗粒的摩尔质量估计为4，640 kg/ mol。该疫苗颗粒在80 ℃以下的温度下是稳定的。组分多肽还显示出在SDS存在下对加热的稳定性。通过在游离溶液中的电泳光散射，估计粒子的等电点为4.0至4.2。希望这些结果和方法可以发挥作用，为未来的表征疫苗颗粒有望在不久的将来开发利用基因工程技术。

Vaccines superior to the conventional ones can now be produced in principle using gene engineering technique. Human hepatitis B virus surface antigen vaccine derived from yeast(y-HBsAg vaccine)is, however, the first and only successful outcome of the introduction of the engineering technique into vaccine production. Its efficacy is believed to be derived from the particular nature and high stability of the assembly composed mainly of protein polypeptides and lipids. The present research project was motivated from our belief that the mode of assembly of the constituents and the source of the stability must be understood to provide basic knowledge hopefully leading to production other efficient vaccines through the approach.The y-HBsAg vaccine particles were found to be spherical, and homogeneous with respect to their size having radius of about 19 nm as judged from dynamic light scattering, small-angle X-ray scattering and electron microscopy. No regular structure was detected in the vaccine particle by small-angle X-ray scattering. The vaccine particle was eluted from a TSK-Gel G600OPW column as a symmetrical peak. Monitoring of the elution gave molar mass estimate of 5, 040 kg/mol for the vaccine particle of the target in the present study of which constituent polypeptide has 9 additional amino acid residues in the N-terminal side compared to the polypeptide of the virus surface antigen. The molar mass of the vaccine particle consisted of the latter polypeptide was estimated to be 4, 640 kg/ mol. The vaccine particle was stable against heating up to 8OﾟC. The constituent polypeptide also showed stability against heating in the presence of SDS. Isoelectric point of the particle was estimated to be 4.0 to 4.2 by electrophoretic light scattering in free solution. It is hoped that these results and approaches can play a role of guidepost for future characterization of vaccine particles expected to be developed in near future using gene engineering technique.