Neutralizing Epitopes in the Membrane-Proximal External Region of HIV-1 gp41 Are Influenced by the Transmembrane Domain and the Plasma Membrane
Neutralizing Epitopes in the Membrane-Proximal External Region of HIV-1 gp41 Are Influenced by the Transmembrane Domain and the Plasma Membrane
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DOI：
10.1128/jvi.06349-11
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发表时间：
2012-03-01
影响因子：
5.4
通讯作者：
Scott, Jamie K.
中科院分区：
文献类型：
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作者：
Montero, Marinieve;Gulzar, Naveed;Scott, Jamie K.
针对HIV-1 gp 41（MPER）的近膜外部区域未能引发广泛中和（bNt）抗体（Abs）反映了模拟其中和活性结构（NCS）的困难。在这里，我们分析了MPER抗原性的质膜的情况下，并确定了gp 41跨膜结构域（TM）在暴露的表位的三个bNt单克隆抗体（单克隆抗体）（2F 5，4 E10，和Z13 e1）的作用。我们瞬时表达了编码gp 41胞外域片段的DNA构建体，该片段与血小板衍生生长因子受体（PDGFR）的TM或gp 41 TM和胞质尾区（CT）融合。编码MPER的构建体与gp 41 TM连接，随后是27个残基的CT片段（MPER-TM 1），产生最佳的MAb结合。使用一组24个MPER-TM 1突变体（在MPER中具有单个氨基酸取代）鉴定了三种Nt MAb的关键结合残基;许多突变体先前显示影响MAb介导的病毒中和。此外，MAb 2F 5和4 E10的非Nt突变体表现出与MPER-TM 1结合的降低，但仍保持与合成MPER肽的结合，表明MPER-TM 1比肽更接近MPER NCS。用PDGFR TM替换MPER-TM 1的gp 41 TM和CT减少了MAb 4 E10的结合，但不减少2F 5的结合，表明gp 41 TM在定向4 E10表位中起关键作用，并且更全面地影响MPER暴露。
Failure to elicit broadly neutralizing (bNt) antibodies (Abs) against the membrane-proximal external region of HIV-1 gp41 (MPER) reflects the difficulty of mimicking its neutralization-competent structure (NCS). Here, we analyzed MPER antigenicity in the context of the plasma membrane and identified a role for the gp41 transmembrane domain (TM) in exposing the epitopes of three bNt monoclonal Abs (MAbs) (2F5, 4E10, and Z13e1). We transiently expressed DNA constructs encoding gp41 ectodomain fragments fused to either the TM of the platelet-derived growth factor receptor (PDGFR) or the gp41 TM and cytoplasmic tail domain (CT). Constructs encoding the MPER tethered to the gp41 TM followed by a 27-residue CT fragment (MPER-TM1) produced optimal MAb binding. Critical binding residues for the three Nt MAbs were identified using a panel of 24 MPER-TM1 mutants bearing single amino acid substitutions in the MPER; many were previously shown to affect MAb-mediated viral neutralization. Moreover, non-Nt mutants of MAbs 2F5 and 4E10 exhibited a reduction in binding to MPER-TM1 and yet maintained binding to synthetic MPER peptides, indicating that MPER-TM1 better approximates the MPER NCS than peptides. Replacement of the gp41 TM and CT of MPER-TM1 with the PDGFR TM reduced binding by MAb 4E10, but not 2F5, indicating that the gp41 TM plays a pivotal role in orienting the 4E10 epitope, and more globally, in affecting MPER exposure.