Choulier L, Laune D, Orfanoudakis G, Wlad H, Janson J, Granier C, Altschuh D
UPR9021-CNRS, IBMC, 15 rue Descartes, 67084 Cedex, Strasbourg, France.
J Immunol Methods. 2001 Mar 1;249(1-2):253-64. doi: 10.1016/s0022-1759(00)00358-6.
Two different approaches, the phage display technique and the Spot peptide synthesis on cellulose membranes, were used to identify sequences recognized by Fab 57P, specific for tobacco mosaic virus protein (TMVP), and define the preferred chemical composition of a functional epitope. Kinetic measurements of the interaction between peptide variants and the antibody fragment were used to further refine the molecular basis of binding activity. Our results show that the functional epitope of Fab 57P requires precise physico-chemical properties at a limited number of positions, and that residues flanking these key residues can influence binding affinity. The phage display and Spot synthesis methods allowed the straightforward localization of the binding region and the identification of residues that are essential for recognition. However, these methods yielded slightly different views of accessory factors that are able to influence antibody binding. The influence on binding activity of these factors can only be assessed through quantitative affinity measurements.
采用两种不同方法，即噬菌体展示技术和纤维素膜上的斑点肽合成法，来鉴定烟草花叶病毒蛋白（TMVP）特异性的Fab 57P所识别的序列，并确定功能性表位的优选化学组成。利用肽变体与抗体片段之间相互作用的动力学测量来进一步完善结合活性的分子基础。我们的结果表明，Fab 57P的功能性表位在有限数量的位置需要精确的物理化学性质，并且这些关键残基侧翼的残基可以影响结合亲和力。噬菌体展示和斑点合成方法能够直接定位结合区域并鉴定识别所必需的残基。然而，这些方法对能够影响抗体结合的辅助因子给出了略有不同的观点。这些因子对结合活性的影响只能通过定量亲和力测量来评估。
