Antigenic topology of the P29 surface lipoprotein of Mycoplasma fermentans: differential display of epitopes results in high-frequency phase variation.
Antigenic topology of the P29 surface lipoprotein of Mycoplasma fermentans: differential display of epitopes results in high-frequency phase variation.
复制标题
发酵支原体 P29 表面脂蛋白的抗原拓扑：表位的差异显示导致高频相位变化。
DOI：
10.1128/iai.64.5.1800-1809.1996
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发表时间：
1996
影响因子：
3.1
通讯作者：
Wise,KS
中科院分区：
文献类型：
--
作者：
Theiss,P;Karpas,A;Wise,KS
P29是发酵支原体的一种主要脂质修饰的表面蛋白，其抗体揭示了在生物体的克隆谱系中以高频率发生的表面表位的相位变化。尽管整个表位携带P29产物的连续表达（通过Western免疫印迹检测），但仍发生这种情况，并与由蛋白质差异表达介导的其他表面抗原的相位变化形成对比。为了了解P29的结构和抗原拓扑学，从菌株PG 18中克隆了单拷贝p29基因并测序。该基因编码含有信号序列的前脂蛋白，该信号序列预测被脂质修饰并在成熟P29脂蛋白的N-末端Cys-1残基处裂解。预测P29的剩余218个残基的亲水性序列位于单个质膜的外部。在成熟蛋白中的位置91和128处的另外的Cys残基显示通过选择性标记仅在单体P29的化学还原后释放的巯基形成36个残基的二硫环。两个几乎相同的带电氨基酸序列出现在P29中，在二硫环内和该结构的上游。通过对PCR产生的p29基因3'缺失突变在大肠杆菌中表达的产物作图，将结合不同单克隆抗体的两个不同表位与P29蛋白的相对末端相关联。每种单克隆抗体检测到高频率和非协调的变化，在克隆变体的菌落免疫印迹中的相应表位的可及性，但从这些变体的p29基因的测序和二硫键的分析显示没有相关的变化，在一级序列或二硫环结构的P29。这些结果表明，P29表面表位的变化可能涉及掩蔽选定的区域的P29，可能是由其他表面组件进行相位变化的差异表达。差异掩蔽可能是改变这种支原体和其他无壁支原体的抗原或功能表面拓扑结构的重要机制。
Antibodies to P29, a major lipid-modified surface protein of Mycoplasma fermentans, reveal phase variation of surface epitopes occurring with high frequency in clonal lineages of the organism. This occurs despite continuous expression of the entire epitope-bearing P29 product (detected by Western immunoblotting) and contrasts with phase variation of other surface antigens mediated by differential expression of proteins. To understand the structure and antigenic topology of P29, the single-copy p29 gene from strain PG18 was cloned and sequenced. The gene encodes a prolipoprotein containing a signal sequence predicted to be modified with lipid and cleaved at the N-terminal Cys-1 residue of the mature P29 lipoprotein. The remaining 218-residue hydrophilic sequence of P29 is predicted to be located external to the single plasma membrane. Additional Cys residues at positions 91 and 128 in the mature protein were shown to form a 36-residue disulfide loop by selectively labeling sulfhydryl groups that were liberated only after chemical reduction of monomeric P29. Two nearly identical charged amino acid sequences occurred in P29, within the disulfide loop and upstream of this structure. Two distinct epitopes binding different monoclonal antibodies were associated with opposite ends of the P29 protein, by mapping products expressed in Escherichia coli from PCR-generated 3' deletion mutations of the p29 gene. Each monoclonal antibody detected high-frequency and noncoordinate changes in accessibility of the corresponding epitopes in colony immunoblots of clonal variants, yet sequencing of the p29 gene from these variants and analysis of disulfide bonds revealed no associated changes in the primary sequence or disulfide loop structure of P29. These results suggest that P29 surface epitope variation may involve masking of selected regions of P29, possibly by other surface components undergoing phase variation by differential expression. Differential masking may be an important mechanism for altering the antigenic or functional surface topology of this mycoplasma and other wall-less mycoplasmas.