Luo Tian, Patel Jignesh G, Zhang Xiaofeng, Walker David H, McBride Jere W
Department of Pathology, University of Texas Medical Branch, Galveston, TX USA.
Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, TX USA.
NPJ Vaccines. 2020 Sep 11;5:85. doi: 10.1038/s41541-020-00231-1. eCollection 2020.
Immunomolecular characterization of (.) and (.) has defined protein orthologs, including tandem repeat proteins (TRPs) that have immunodominant linear antibody epitopes. In this study, we combined bioinformatic analysis and cell-free protein expression to identify undiscovered immunoreactive . and . hypothetical proteins. Antigenicity of the . and . ORFeomes ( = 1105 and = 925, respectively) was analyzed by the sequence-based prediction model ANTIGENpro, and we identified ~250 ORFs in each respective ORFeome as highly antigenic. The hypothetical proteins (. = 93 and . = 98) present in the top 250 antigenic ORFs were further investigated in this study. By ELISA, 46 . and 30 . IVTT-expressed hypothetical proteins reacted with antibodies in sera from naturally .-infected patients or .-infected dogs. Moreover, 15 . and 16 . proteins consistently reacted with a panel of sera from patients or dogs, including many that revealed the immunoreactivity of "gold standard" TRPs. Antibody epitopes in most (>70%) of these proteins exhibited partial or complete conformation-dependence. The majority (23/31; 74%) of the major immunoreactive proteins identified were small (≤250 aa), and 20/31 (65%) were predicted to be secreted effectors. Unlike the strong linear antibody epitopes previously identified in TRP and OMP orthologs, there were contrasting differences in the . and . antigenic repertoires, epitopes and ortholog immunoreactivity. This study reveals numerous previously undefined immunodominant and subdominant antigens, and illustrates the breadth, complexity, and diversity of immunoreactive proteins/epitopes in .
（.）和（.）的免疫分子特征已确定了蛋白质直系同源物，包括具有免疫显性线性抗体表位的串联重复蛋白（TRP）。在本研究中，我们结合生物信息学分析和无细胞蛋白质表达来鉴定未发现的免疫反应性.和.假设蛋白。通过基于序列的预测模型ANTIGENpro分析了.和.开放阅读框组（分别为1105个和925个）的抗原性，我们在每个开放阅读框组中鉴定出约250个开放阅读框为高抗原性。本研究进一步调查了前250个抗原性开放阅读框中存在的假设蛋白（.为93个，.为98个）。通过酶联免疫吸附测定（ELISA），46个.和30个.体外转录翻译（IVTT）表达的假设蛋白与自然感染.的患者或感染.的犬血清中的抗体发生反应。此外，15个.和16个.蛋白与一组患者或犬的血清持续发生反应，其中许多血清显示出“金标准”TRP的免疫反应性。这些蛋白中大多数（>70%）的抗体表位表现出部分或完全的构象依赖性。鉴定出的主要免疫反应性蛋白中的大多数（23/31；74%）较小（≤250个氨基酸），并且20/31（65%）被预测为分泌效应子。与先前在TRP和外膜蛋白（OMP）直系同源物中鉴定出的强线性抗体表位不同，.和.的抗原库、表位和直系同源物免疫反应性存在显著差异。本研究揭示了许多先前未定义的免疫显性和亚显性抗原，并说明了.中免疫反应性蛋白/表位的广度、复杂性和多样性。
