Analysis of the Mycoplasma genitalium MgpB Adhesin to Predict Membrane Topology, Investigate Antibody Accessibility, Characterize Amino Acid Diversity, and Identify Functional and Immunogenic Epitopes

Mycoplasma genitalium is a sexually transmitted pathogen and is associated with reproductive tract disease that can be chronic in nature despite the induction of a strong antibody response. Persistent infection exacerbates the likelihood of transmission, increases the risk of ascension to the upper tract, and suggests that M. genitalium may possess immune evasion mechanism(s). Antibodies from infected patients predominantly target the MgpB adhesin, which is encoded by a gene that recombines with homologous donor sequences, thereby generating sequence variation within and among strains. We have previously characterized mgpB heterogeneity over the course of persistent infection and have correlated the induction of variant-specific antibodies with the loss of that particular variant from the infected host. In the current study, we examined the membrane topology, antibody accessibility, distribution of amino acid diversity, and the location of functional and antigenic epitopes within the MgpB adhesin. Our results indicate that MgpB contains a single transmembrane domain, that the majority of the protein is surface exposed and antibody accessible, and that the attachment domain is located within the extracellular C-terminus. Not unexpectedly, amino acid diversity was concentrated within and around the three previously defined variable regions (B, EF, and G) of MgpB; while nonsynonymous mutations were twice as frequent as synonymous mutations in regions B and G, region EF had equal numbers of nonsynonymous and synonymous mutations. Interestingly, antibodies produced during persistent infection reacted predominantly with the conserved C-terminus and variable region B. In contrast, infection-induced antibodies reacted poorly with the N-terminus, variable regions EF and G, and intervening conserved regions despite the presence of predicted B cell epitopes. Overall, this study provides an important foundation to define how different segments of the MgpB adhesin contribute to functionality, variability, and immunogenicity during persistent M. genitalium infection.

生殖支原体（Mycoplasma genitalium）是一种性传播病原体，与生殖道疾病密切相关；尽管机体可诱导强烈的抗体应答，但该类疾病仍可呈慢性病程。持续感染会增加病原体的传播风险，提升其上行至上生殖道的概率，同时提示生殖支原体可能具备免疫逃逸机制（immune evasion mechanism(s)）。

感染患者体内的抗体主要靶向MgpB黏附素（MgpB adhesin），该黏附素由可与同源供体序列（homologous donor sequences）发生重组的基因编码，由此可在菌株内部及不同菌株间产生序列变异。我们此前已对持续感染过程中的mgpB异质性进行了表征，并将变异株特异性抗体的诱导与该特定变异株从感染宿主体内的清除建立了关联。

在本研究中，我们对MgpB黏附素的膜拓扑结构（membrane topology）、抗体可及性（antibody accessibility）、氨基酸多样性分布，以及功能与抗原表位的位置进行了系统分析。研究结果显示，MgpB仅含一个跨膜结构域（transmembrane domain），该蛋白的大部分区域暴露于菌体表面且可与抗体结合，其附着结构域（attachment domain）定位于细胞外C端区域。

不出所料，氨基酸多态性集中于MgpB此前定义的三个可变区（B、EF和G）及其周边区域；在B区和G区，非同义突变（nonsynonymous mutations）的发生频率为同义突变（synonymous mutations）的两倍，而EF区的两类突变数量相当。有趣的是，持续感染过程中产生的抗体主要与保守C端及可变区B发生反应。与之相反，尽管存在预测的B细胞表位（B cell epitopes），但感染诱导产生的抗体与N端、可变区EF和G以及中间的保守区域的结合活性较弱。

总体而言，本研究为阐明生殖支原体持续感染过程中，MgpB黏附素的不同区段如何影响其功能、变异性及免疫原性（immunogenicity）提供了重要的研究基础。