Abstract：
BACKGROUND: Pneumocystis jirovecii is the most emerging life-threating health problem that causes acute and fatal pneumonia infection. It is rare and more contagious for patients with leukemia and immune-deficiency disorders. Until now there is no treatment available for this infection therefore, it is needed to develop any treatment against this pathogen.
METHODS: In this work, we used comparative proteomics, robust immune-informatics, and reverse vaccinology to create an mRNA vaccine against Pneumocystis jirovecii by targeting outer and transmembrane proteins. Using a comparative subtractive proteomic analysis of two Pneumocystis jirovecii proteomes, a distinct non-redundant Pneumocystis jirovecii (strain SE8) proteome was chosen. Seven Pneumocystis jirovecii transmembrane proteins were chosen from this proteome based on hydrophilicity, essentiality, virulence, antigenicity, pathway interaction, protein-protein network analysis, and allergenicity.
OBJECTIVE: The reverse vaccinology approach was used to predict the immunogenic and antigenic epitopes of major histocompatibility complex (MHC) I, II and B-cells from the selected proteins on the basis of their antigenicity, toxicity and allergenicity. These immunogenic epitopes were linked together to construct the mRNA-based vaccine. To enhance the immunogenicity, suitable adjuvant, linkers (GPGPG, KK, and CYY), and PRDRE sequences were used.
RESULTS: Through predictive modeling and confirmation via the Ramachandran plot, we assessed secondary and 3D structures. The adjuvant RpfE was incorporated to enhance the vaccine construct\'s immunogenicity (GRAVY index: -0.271, instability index: 39.53, antigenicity: 1.0428). The physiochemical profiling of vaccine construct was predicted it an antigenic, efficient, and potential vaccine. Notably, strong interactions were observed between the vaccine construct and TLR-3/TLR-4 (-1301.7 kcal/mol-1 and -1374.7 kcal/mol-1).
CONCLUSIONS: The results predicted that mRNA-based vaccines trigger a cellular and humoral immune response, making the vaccine potential candidate against Pneumocystis jirovecii and it is more suitable for in-vitro analysis and validation to prove its effectiveness.
METHODS: In this work, we used comparative proteomics, robust immune-informatics, and reverse vaccinology to create an mRNA vaccine against Pneumocystis jirovecii by targeting outer and transmembrane proteins. Using a comparative subtractive proteomic analysis of two Pneumocystis jirovecii proteomes, a distinct non-redundant Pneumocystis jirovecii (strain SE8) proteome was chosen. Seven Pneumocystis jirovecii transmembrane proteins were chosen from this proteome based on hydrophilicity, essentiality, virulence, antigenicity, pathway interaction, protein-protein network analysis, and allergenicity.
OBJECTIVE: The reverse vaccinology approach was used to predict the immunogenic and antigenic epitopes of major histocompatibility complex (MHC) I, II and B-cells from the selected proteins on the basis of their antigenicity, toxicity and allergenicity. These immunogenic epitopes were linked together to construct the mRNA-based vaccine. To enhance the immunogenicity, suitable adjuvant, linkers (GPGPG, KK, and CYY), and PRDRE sequences were used.
RESULTS: Through predictive modeling and confirmation via the Ramachandran plot, we assessed secondary and 3D structures. The adjuvant RpfE was incorporated to enhance the vaccine construct\'s immunogenicity (GRAVY index: -0.271, instability index: 39.53, antigenicity: 1.0428). The physiochemical profiling of vaccine construct was predicted it an antigenic, efficient, and potential vaccine. Notably, strong interactions were observed between the vaccine construct and TLR-3/TLR-4 (-1301.7 kcal/mol-1 and -1374.7 kcal/mol-1).
CONCLUSIONS: The results predicted that mRNA-based vaccines trigger a cellular and humoral immune response, making the vaccine potential candidate against Pneumocystis jirovecii and it is more suitable for in-vitro analysis and validation to prove its effectiveness.
摘要：
背景：jirovecii肺孢子虫是引起急性和致命性肺炎感染的最新出现的危及生命的健康问题。对于患有白血病和免疫缺陷疾病的患者来说，这是罕见的，更具传染性。因此，到目前为止还没有治疗这种感染的方法，需要开发针对这种病原体的任何治疗方法。
方法：在这项工作中，我们使用了比较蛋白质组学，强大的免疫信息学，和反向疫苗学，通过靶向外膜蛋白和跨膜蛋白来创建针对jirovecii肺孢子虫的mRNA疫苗。使用两种肺孢子虫蛋白质组的比较消减蛋白质组学分析，选择了不同的非冗余肺孢子虫（菌株SE8）蛋白质组。基于亲水性，从该蛋白质组中选择了七个jirovecii肺孢子虫跨膜蛋白，本质，毒力,抗原性,途径相互作用，蛋白质-蛋白质网络分析，和过敏原性。
目的：反向疫苗学方法用于预测主要组织相容性复合体（MHC）I的免疫原性和抗原表位，II和B细胞从选定的蛋白质的基础上，它们的抗原性，毒性和致敏性。将这些免疫原性表位连接在一起以构建基于mRNA的疫苗。为了增强免疫原性，合适的佐剂，接头（GPGPG，KK,和CYY），和PRDRE序列被使用。
结果：通过Ramachandran图的预测建模和确认，我们评估了二级和三维结构。掺入佐剂RpfE以增强疫苗构建体的免疫原性(GRAVY指数:-0.271,不稳定性指数:39.53,抗原性:1.0428)。疫苗构建体的理化分析被预测为抗原性，高效,和潜在的疫苗。值得注意的是,在疫苗构建体和TLR-3/TLR-4(-1301.7kcal/mol-1和-1374.7kcal/mol-1)之间观察到强相互作用。
结论：结果预测基于mRNA的疫苗会引发细胞和体液免疫反应，使疫苗成为对抗jirovecii肺孢子虫的潜在候选物，并且更适合用于体外分析和验证以证明其有效性。
方法：在这项工作中，我们使用了比较蛋白质组学，强大的免疫信息学，和反向疫苗学，通过靶向外膜蛋白和跨膜蛋白来创建针对jirovecii肺孢子虫的mRNA疫苗。使用两种肺孢子虫蛋白质组的比较消减蛋白质组学分析，选择了不同的非冗余肺孢子虫（菌株SE8）蛋白质组。基于亲水性，从该蛋白质组中选择了七个jirovecii肺孢子虫跨膜蛋白，本质，毒力,抗原性,途径相互作用，蛋白质-蛋白质网络分析，和过敏原性。
目的：反向疫苗学方法用于预测主要组织相容性复合体（MHC）I的免疫原性和抗原表位，II和B细胞从选定的蛋白质的基础上，它们的抗原性，毒性和致敏性。将这些免疫原性表位连接在一起以构建基于mRNA的疫苗。为了增强免疫原性，合适的佐剂，接头（GPGPG，KK,和CYY），和PRDRE序列被使用。
结果：通过Ramachandran图的预测建模和确认，我们评估了二级和三维结构。掺入佐剂RpfE以增强疫苗构建体的免疫原性(GRAVY指数:-0.271,不稳定性指数:39.53,抗原性:1.0428)。疫苗构建体的理化分析被预测为抗原性，高效,和潜在的疫苗。值得注意的是,在疫苗构建体和TLR-3/TLR-4(-1301.7kcal/mol-1和-1374.7kcal/mol-1)之间观察到强相互作用。
结论：结果预测基于mRNA的疫苗会引发细胞和体液免疫反应，使疫苗成为对抗jirovecii肺孢子虫的潜在候选物，并且更适合用于体外分析和验证以证明其有效性。
