Kay B K, Adey N B, He Y S, Manfredi J P, Mataragnon A H, Fowlkes D M
Department of Biology, University of North Carolina, Chapel Hill 27599-3280.
Gene. 1993 Jun 15;128(1):59-65. doi: 10.1016/0378-1119(93)90153-t.
We have examined the potential of isolating novel ligands from a library of M13 pIII-fusion phage displaying peptides composed of 38 random amino acids (aa). The library was panned with streptavidin (SA) and a polyclonal goat antimouse IgG Fc antibody (Ab) preparation coupled to paramagnetic beads. SA selected two classes of phage from the library. One class exhibited the aa motif, HP(Q/M) theta (where theta signifies a non-polar aa), similar to the motif identified by Devlin et al. [Science 249 (1990) 404-406] using a 15-aa random peptide library displayed on phage. The other class of phage had no discernible motif. In binding experiments, the non-HP(Q/M) theta phage had a slightly higher affinity for SA than did the motif phage. Both classes of SA-binding phage failed to bind native and non-glycosylated forms of avidin, even though SA and avidin are structurally similar and both proteins possess extraordinary affinities for biotin. The polyclonal goat anti-mouse IgG Fc Ab preparation selected phage displaying sequences similar to a region of the mouse IgG Fc. Thus, a single immunodominant epitope on the mouse IgG Fc was identified. Furthermore, a second phage displaying peptides with no discernible sequence similarities to mouse IgG Fc was isolated. Thus, an M13 library displaying 38-aa peptides can yield phage with affinity for various targets. Finally, we have observed a biological bias against odd numbers of Cys residues in the displayed peptides.
我们研究了从一个M13 pIII融合噬菌体文库中分离新型配体的潜力，该文库展示由38个随机氨基酸（aa）组成的肽。该文库用链霉亲和素（SA）和与顺磁性珠偶联的多克隆山羊抗小鼠IgG Fc抗体（Ab）制剂进行淘选。SA从文库中筛选出两类噬菌体。一类呈现aa基序HP(Q/M)θ（其中θ表示非极性aa），类似于Devlin等人[《科学》249 (1990) 404 - 406]使用展示在噬菌体上的15个aa随机肽文库鉴定出的基序。另一类噬菌体没有可识别的基序。在结合实验中，非HP(Q/M)θ噬菌体对SA的亲和力略高于基序噬菌体。尽管SA和抗生物素蛋白在结构上相似且两种蛋白质对生物素都具有非凡的亲和力，但这两类与SA结合的噬菌体都不能结合天然和非糖基化形式的抗生物素蛋白。多克隆山羊抗小鼠IgG Fc Ab制剂筛选出展示与小鼠IgG Fc区域相似序列的噬菌体。因此，鉴定出了小鼠IgG Fc上的一个单一免疫显性表位。此外，还分离出了第二种展示与小鼠IgG Fc没有明显序列相似性的肽的噬菌体。因此，展示38个aa肽的M13文库可以产生对各种靶标具有亲和力的噬菌体。最后，我们观察到所展示的肽中存在对奇数个半胱氨酸残基的生物学偏好。
