Rheumatoid-factor-reactive sites on CH3 established by overlapping 7-mer peptide epitope analysis.
Rheumatoid-factor-reactive sites on CH3 established by overlapping 7-mer peptide epitope analysis.
复制标题
通过重叠 7 聚体肽表位分析建立 CH3 上的类风湿因子反应位点。
DOI：
10.1016/0161-5890(94)00122-h
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发表时间：
1995
影响因子：
3.6
通讯作者：
WilliamsJr,RC
中科院分区：
文献类型：
--
作者：
Peterson,C;Malone,CC;WilliamsJr,RC
从10例类风湿性关节炎患者和6个单克隆IgM类风湿因子（RF）的多克隆IgM RF分离的单体IgG亲和柱，并研究其与整个CH 3结构域的IgG合成为重叠的7-聚体使用pin-ELISA测定的反应性。所有10个多克隆IgM RF显示出相似的反应性特征，包括具有溶剂可及残基PREPQVY的肽（残基343-349），PQVYTLP（残基346-352）、TLPPRSE（350-356）、DGSFFLY（401-407）、WQQGNVF（417-423）、CSVMHEG（425-430）、EGLHNHY（430-436）和KSLSLSP（439-446）。用中性甘氨酸或丙氨酸取代这些RF反应性表位中的每个残基表明，349位的酪氨酸，343、346和352位的脯氨酸，谷氨酰胺347，缬氨酸348，苏氨酸350，亮氨酸351，精氨酸354，天冬氨酸401，酪氨酸407，丝氨酸426，组氨酸429，亮氨酸432，酪氨酸436和赖氨酸439代表CH 3中RF反应性的重要单个氨基酸。CH 3一级序列中含有或不含有甘氨酸取代丙氨酸431（Gmx）或天冬氨酸取代谷氨酸（356）和亮氨酸取代甲硫氨酸（358）（Gma）的单一同种异型特异性氨基酸取代的区域，与单个多克隆和单克隆RF的反应性通常显示出相当大的差异。然而，RF反应性的这些差异与个体抗Gm RF特异性无关。使用从RA滑膜B细胞或外周血B细胞产生的单克隆IgM RF的测定经常显示出反应性CH 3表位的更有限的谱。7-代表与多克隆IgM RF预孵育的CH 3上的RF反应位点的mer肽显示出对ELISA板上RF与整个IgG结合的强烈抑制（55-66%）。这些研究表明，它是可能的，以确定部分的IgG CH 3结构域参与反应与IgM RF使用反应性表位映射与连续的线性肽衍生的主要IgG CH 3序列。
Polyclonal IgM rheumatoid factors (RF) from ten patients with rheumatoid arthritis and six monoclonal IgM RF were isolated from monomeric IgG affinity columns and studied for their reactivity with the entire CH3domain of IgG synthesized as overlapping 7-mers using a pin-ELISA assay. All ten polyclonal IgM RF showed similar profiles of reactivity which included peptides with solvent accessible residues PREPQVY (residues 343–349), PQVYTLP (residues 346–352), TLPPRSE (350–356), DGSFFLY (401–407), WQQGNVF (417–423), CSVMHEG (425–430), EGLHNHY (430–436) and KSLSLSP (439–446) of the CH3domain. Substitution of a neutral glycine or alanine for each residue within these RF-reactive epitopes indicated that tyrosine at position 349, prolines at 343, 346 and 352, glutamine 347, valine 348, threonine 350, leucine 351, arginine 354, aspartic acid 401, tyrosine 407, serine 426, histidine 429, leucine 432, tyrosine 436 and lysine 439 represented important single amino acids within CH3for RF reactivity. Regions of CH3primary sequence with and without the single allotype-specific amino acid substitutions of glycine for alanine 431 (Gmx) or aspartic acid for glutamic acid (356) and leucine for methionine (358) (Gma) often showed considerable differences in reactivity with individual polyclonal and monoclonal RF. However, these differences in RF reactivity did not correlate with the individual anti-Gm RF specificity. Assays using monoclonal IgM RF produced from RA synovial B cells or peripheral blood B cells frequently showed a much more restricted spectrum of reactive CH3epitopes. 7-mer peptides representing RF-reactive sites on CH3preincubated with polyclonal IgM RF showed strong inhibition (55–66%) of RF binding to whole IgG on the ELISA plate. These studies indicate that it is possible to define portions of the IgG CH3domain participating in the reaction with IgM RF using reactive epitope-mapping with sequential linear peptides derived from the primary IgG CH3sequence.