Hudson N W, Albrecht M, Randhawa Z I, Steinrauf L K, Hamilton J A
Northwest Center for Medical Education, Indiana University School of Medicine, Gary 46408.
Mol Immunol. 1993 May;30(7):685-93. doi: 10.1016/0161-5890(93)90080-u.
A panel of antibodies which differ in their L chain structures and which bind to structurally defined haptens, would be useful in investigating L chain structure and function. In a previous study, chain recombinant antibody CR24 (26-10 H, 45-20 lambda) was produced by hybridoma-hybridoma fusion. Although both parental antibodies bound digoxin with high affinity, CR24 lacked detectable digoxin-binding activity. Hybridoma CR24 was subsequently fused with H chain-loss hybridomas in order to produce a panel of antibodies composed of 26-10 H chains and 26-10 "like" L chains. Two antibodies produced were CR260 which demonstrated digoxin-binding activity and CR256 which did not. CR260 and CR256 expressed only one amino acid difference (Pro to Leu at L-96). This difference resulted in the CR256 binding defect. In this report, two new antidigoxin antibodies are described. One, SR2E7, contained the Pro to Leu (L-96) defect, but still bound digoxin. Binding affinities and binding specificity patterns, as well as complete VL DNA sequence and corresponding protein sequence of the new digoxin binding antibody L chains (SR2E7 and SR1C7) are presented. Both kappa L chains are highly homologous to the 26-10 kappa L chain as well as the BALB/c germline gene K5.1. These results suggest that antibodies which are initially defective in binding activity can be cured by changing specific amino acids involved in determining the binding-site structure. Molecular modelling studies of the binding-site region were completed to address L chain structural changes induced by specific amino acid substitutions.
一组轻链结构不同且能与结构明确的半抗原结合的抗体，将有助于研究轻链的结构和功能。在先前的一项研究中，通过杂交瘤-杂交瘤融合产生了链重组抗体CR24（26-10 H，45-20 λ）。尽管两种亲本抗体都以高亲和力结合地高辛，但CR24缺乏可检测到的地高辛结合活性。随后，杂交瘤CR24与重链缺失杂交瘤融合，以产生一组由26-10重链和26-10“类”轻链组成的抗体。产生的两种抗体是具有地高辛结合活性的CR260和不具有该活性的CR256。CR260和CR256仅表达一个氨基酸差异（轻链96位脯氨酸突变为亮氨酸）。这种差异导致了CR256的结合缺陷。在本报告中，描述了两种新的抗地高辛抗体。一种是SR2E7，具有脯氨酸到亮氨酸（轻链96位）的缺陷，但仍能结合地高辛。本文给出了新的地高辛结合抗体轻链（SR2E7和SR1C7）的结合亲和力和结合特异性模式，以及完整的VL DNA序列和相应的蛋白质序列。两种κ轻链与26-10 κ轻链以及BALB/c种系基因K5.1高度同源。这些结果表明，最初结合活性有缺陷的抗体可以通过改变参与确定结合位点结构的特定氨基酸来修复。完成了结合位点区域的分子建模研究，以探讨特定氨基酸取代引起的轻链结构变化。
