Research Center for Biotechnology, Research Organization of Life Sciences, National Research and Innovation Agency of The Republic of Indonesia (BRIN), Cibinong Sciences Center, Cibinong, Bogor, Indonesia.
Prep Biochem Biotechnol. 2023;53(2):148-156. doi: 10.1080/10826068.2022.2048262. Epub 2022 Mar 18.
Cervical cancer caused by (HPV) is one of the most common causes of cancer death in women worldwide. Even though the disease can be avoided by immunization, the expensive price of HPV vaccines makes it hard to be accessed by women in middle-low-income countries. Thus, the development of generic HPV vaccines is needed to address inequalities in life-saving access. This study aimed to develop the HPV52 L1 VLP-based recombinant vaccine using expression system. The gene was codon-optimized based on codon usage resulting CAI value of 0.804. The gene was inserted into the pD902 plasmid under the regulation of the promoter. The linear plasmid was transformed into BG10 genome and screened in YPD medium containing zeocin antibiotic. Colony of transformant that grown on highest zeocin concentration was characterized by genomic PCR and sequencing. The positive clone was selected and expressed using BMGY/BMMY medium induced with various methanol concentrations. The SDS-PAGE and Western blot analyses showed that 55 kDa L1 protein was successfully expressed using an optimum concentration of 1% methanol. The self-assembly of HPV52 L1 protein was also proven using TEM analysis. Moreover, we also analyzed the B-cell epitope of HPV52 L1 protein based on several criteria, including antigenicity, surface accessibility, flexibility, and hydrophilicity. We assumed that epitope GLQARPKLKRPASSAPRTSTKKKKV could be developed as an epitope-based vaccine with a neutralizing antibody response toward HPV52 infection. Finally, our study provided the alternative for developing low-cost HPV vaccines, either VLP or epitope-based.
由 （HPV）引起的宫颈癌是全球女性癌症死亡的最常见原因之一。尽管可以通过免疫接种来预防这种疾病，但 HPV 疫苗价格昂贵，使得中低收入国家的妇女难以获得。因此，需要开发通用 HPV 疫苗来解决救生通道方面的不平等问题。本研究旨在使用 表达系统开发基于 HPV52 L1 VLP 的重组疫苗。根据 密码子使用情况，对 基因进行了密码子优化，CAI 值为 0.804。该基因被插入 pD902 质粒中，由 启动子调控。线性质粒转化到 BG10 基因组中，并在含有博来霉素抗生素的 YPD 培养基中筛选。在最高博来霉素浓度下生长的转化体菌落通过基因组 PCR 和测序进行表征。选择阳性克隆，并使用 BMGY/BMMY 培养基在不同甲醇浓度下诱导表达。SDS-PAGE 和 Western blot 分析表明，使用 1%甲醇的最佳浓度可成功表达 55 kDa 的 L1 蛋白。使用 TEM 分析还证明了 HPV52 L1 蛋白的自我组装。此外，我们还根据抗原性、表面可及性、灵活性和亲水性等标准，分析了 HPV52 L1 蛋白的 B 细胞表位。我们假设表位 GLQARPKLKRPASSAPRTSTKKKKV 可以作为一种基于表位的疫苗开发，针对 HPV52 感染产生中和抗体反应。最后，我们的研究为开发低成本 HPV 疫苗（无论是 VLP 还是基于表位的疫苗）提供了替代方案。
