Oldstone M B, Tishon A, Lewicki H, Dyson H J, Feher V A, Assa-Munt N, Wright P E
Department of Neuropharmacology, Research Institute of Scripps Clinic, La Jolla, California 92037.
J Virol. 1991 Apr;65(4):1727-34. doi: 10.1128/JVI.65.4.1727-1734.1991.
Thirty-six monoclonal antibodies from mice and three from rats were raised against a peptide corresponding to the immunodominant domain of the transmembrane gp41 protein of human immunodeficiency virus (HIV) type 1 (LGLWGCSGKLIC; amino acid residues 598 to 609). Of these, three monoclonal antibodies from the mice and one from a rat also reacted with the corresponding peptide derived from the HIV type 2 transmembrane gp41 protein (amino acid residues 593 to 603; NSWGCAFRQVC). Immunochemical studies using a variety of synthetic peptides indicated that the cross-reactivity was due to antibody binding to CSGKLIC of HIV type 1 or CAFRQVC of HIV type 2. Single amino acid substitutions for a cysteine at either the amino or the carboxy end of the peptide interrupted antibody binding, indicating that the site recognized was the Cys-XXXXX-Cys loop. Similar results were obtained when the 11-mer HIV type 2 gp41 peptide (amino acids 593 to 603) was inoculated into mice to raise monoclonal antibodies. In this instance, of 30 monoclonal antibodies developed, 4 reacted with both HIV type 1 and HIV type 2 peptides. The conformation of a seven-residue peptide, CSGKLIC, corresponding to residues 603 to 609 of the gp41 immunodominant epitope of HIV-1 was investigated by proton nuclear magnetic resonance spectroscopy. The immunologically active form of CSGKLIC contains an intramolecular disulfide bond and maintains a preference for a folded conformation, apparently including a type I reverse turn about the residues SGKL. No such preference is observed for the reduced form of the peptide, which contains two thiol groups. The presence of the disulfide bond is thus integral to the formation of the structure of the loop in solution. In agreement with this finding, elimination of the possibility of loop formation by substitution of S for C at the amino or carboxy termini of the 7-mer is accompanied by the failure of antibody binding to this peptide.
针对与人类免疫缺陷病毒1型（HIV-1）跨膜糖蛋白gp41的免疫显性结构域相对应的肽段（LGLWGCSGKLIC；氨基酸残基598至609），制备了36种小鼠单克隆抗体和3种大鼠单克隆抗体。其中，3种小鼠单克隆抗体和1种大鼠单克隆抗体也与源自HIV-2跨膜糖蛋白gp41的相应肽段（氨基酸残基593至603；NSWGCAFRQVC）发生反应。使用多种合成肽进行的免疫化学研究表明，交叉反应性是由于抗体与HIV-1的CSGKLIC或HIV-2的CAFRQVC结合。肽段氨基端或羧基端的半胱氨酸被单个氨基酸取代会中断抗体结合，这表明识别的位点是Cys-XXXXX-Cys环。将11肽HIV-2 gp41肽段（氨基酸593至603）接种到小鼠体内制备单克隆抗体时，也得到了类似结果。在此例中，所产生的30种单克隆抗体中有4种与HIV-1和HIV-2肽段均发生反应。通过质子核磁共振光谱研究了与HIV-1 gp41免疫显性表位的残基603至609相对应的七肽CSGKLIC的构象。CSGKLIC的免疫活性形式包含分子内二硫键，并保持对折叠构象的偏好，显然包括围绕残基SGKL的I型反向转角。对于含有两个巯基的还原形式的肽段，未观察到这种偏好。因此，二硫键的存在对于溶液中环结构的形成是不可或缺的。与此发现一致的是，在七肽的氨基端或羧基端用S取代C消除环形成的可能性，会导致抗体无法与该肽段结合。
