Human immunodeficiency virus type 1-neutralizing monoclonal antibody 2F5 is multispecific for sequences flanking the DKW core epitope.
Human immunodeficiency virus type 1-neutralizing monoclonal antibody 2F5 is multispecific for sequences flanking the DKW core epitope.
复制标题
人类免疫缺陷病毒 1 型中和单克隆抗体 2F5 对 DKW 核心表位侧翼序列具有多特异性。
DOI：
10.1016/j.jmb.2004.02.051
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Scott,JamieK
中科院分区：
文献类型：
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作者：
Menendez,Alfredo;Chow,KeithC;Pan,OscarCC;Scott,JamieK
人单克隆抗体2F 5是少数几种中和广泛的HIV-1原代分离株的人抗体之一。gp 41上的2F 5表位包括序列ELDKWA，其中核心残基DKW对于抗体结合至关重要。用带有ELDKWA的肽免疫从未引起HIV中和抗体，表明2F 5互补位的重要部分仍未鉴定。使用延伸超过ELDKWA的较长肽导致表位抗原性增加，但尚未产生中和抗体。我们试图开发与2F 5结合的肽，并作为2F 5互补位的特异性探针。因此，我们使用2F 5来筛选一组噬菌体展示的随机肽文库。来自随机肽文库的紧密结合克隆在DKW基序侧翼的区域中显示序列变异性。为了进一步揭示参与2F 5结合的侧翼区，构建了两个半限定文库，其具有DKW核心的N-末端或C-末端的12个杂色残基（分别为X12-AADKW和AADKW-X12）。从AADKW-X12文库中分离的三个克隆具有相似的高亲和力，尽管它们之间或与gp 41缺乏序列同源性。通过Ala取代和氨基酸缺失研究评估这些克隆的每个残基对2F 5结合的贡献，并揭示每个克隆通过不同的机制结合2F 5。这些结果表明，2F 5互补位由至少两个功能不同的区域形成：一个显示对DKW核心表位的特异性，另一个对核心表位C端序列具有多特异性。这第二，多特异性区域的2F 5互补位，其独特的生物学功能的影响进行了讨论。
Human monoclonal antibody 2F5 is one of a few human antibodies that neutralize a broad range of HIV-1 primary isolates. The 2F5 epitope on gp41 includes the sequence ELDKWA, with the core residues, DKW, being critical for antibody binding. HIV-neutralizing antibodies have never been elicited by immunization with peptides bearing ELDKWA, suggesting that important part(s) of the 2F5 paratope remain unidentified. The use of longer peptides extending beyond ELDKWA has resulted in increased epitope antigenicity, but neutralizing antibodies have not been generated. We sought to develop peptides that bind to 2F5, and that function as specific probes of the 2F5 paratope. Thus, we used 2F5 to screen a set of phage-displayed, random peptide libraries. Tight-binding clones from the random peptide libraries displayed sequence variability in the regions flanking the DKW motif. To further reveal flanking regions involved in 2F5 binding, two semi-defined libraries were constructed having 12 variegated residues either N-terminal or C-terminal to the DKW core (X12-AADKW and AADKW-X12, respectively). Three clones isolated from the AADKW-X12library had similar high affinities, despite a lack of sequence homology among them, or with gp41. The contribution of each residue of these clones to 2F5 binding was evaluated by Ala substitution and amino acid deletion studies, and revealed that each clone bound 2F5 by a different mechanism. These results suggest that the 2F5 paratope is formed by at least two functionally distinct regions: one that displays specificity for the DKW core epitope, and another that is multispecific for sequences C-terminal to the core epitope. The implications of this second, multispecific region of the 2F5 paratope for its unique biological function are discussed.