Characterization of peptides that bind the tumor-associated Thomsen-Friedenreich antigen selected from bacteriophage display libraries.
Characterization of peptides that bind the tumor-associated Thomsen-Friedenreich antigen selected from bacteriophage display libraries.
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DOI：
10.1006/jmbi.1997.1107
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发表时间：
1997-07
影响因子：
5.6
通讯作者：
E. Peletskaya;V. V. Glinsky-V.;G. Glinsky;S. Deutscher;T. Quinn
中科院分区：
文献类型：
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作者：
E. Peletskaya;V. V. Glinsky-V.;G. Glinsky;S. Deutscher;T. Quinn
先前已经从随机肽噬菌体展示文库中鉴定出对许多蛋白质和核酸具有高亲和力和特异性的肽。在这里，随机肽噬菌体展示文库被用来鉴定与癌症相关的Thomsen-Friedenreich糖抗原（T抗原）结合的序列。存在于大多数恶性细胞上的T抗原含有一种免疫优势的Gal β 1 - bbb3galnac α双糖，在大多数癌细胞表面被发现。这种抗原被认为与肿瘤细胞聚集和转移有关。两个15个氨基酸随机肽噬菌体展示文库与表面显示T抗原的糖蛋白有亲和力。序列分析显示，许多肽与几种碳水化合物结合蛋白的糖识别位点具有同源性。比较两个文库中亲和选择的序列，得到了一个富含芳香氨基酸的共同基序（W-Y-A-W/F-S-P）。根据亲和选择的序列，化学合成了四个肽，并对其碳水化合物识别特性进行了表征。合成的多肽对asialofetuin上显示的T抗原或与牛血清白蛋白结合的T抗原（MAP-P30与asialofetuin结合的Kd = 5 nM）以及溶液中游离的T抗原双糖（MAP-P30的Kd = 10微米，P10的Kd = 20微米）具有较高的特异性和亲和力。P30和P10两种多肽在溶液中对asialofetuin和T抗原均表现出较高的亲和力和特异性。每个序列中单独的酪氨酸残基的碘化显著降低了它们结合T抗原的能力，这表明酪氨酸残基在碳水化合物识别中起着重要作用。P30和P10能够在体外抑制asialofetin介导的黑色素瘤细胞聚集，并与花生凝集素竞争结合MDA-MB-435乳腺癌细胞表面的T抗原，这证明了这些肽具有重要的功能意义。
Peptides with high affinities and specificities for numerous proteins and nucleic acids have been previously identified from random peptide bacteriophage display libraries. Here, random peptide bacteriophage display libraries were used to identify sequences that bound the cancer-associated Thomsen-Friedenreich glycoantigen (T antigen). The T antigen, present on most malignant cells, contains an immunodominant Gal beta1 --> 3GalNAc alpha disaccharide unmasked on the surfaces of most carcinomas. This antigen has been postulated to be involved in tumor cell aggregation and metastasis. Two 15 amino acid random peptide bacteriophage display libraries were affinity selected with glycoproteins displaying T antigen on their surfaces. Sequence analysis revealed that many of the peptides shared homology with sugar recognition sites in several carbohydrate-binding proteins. A comparison of affinity selected sequences from both libraries yielded a common motif (W-Y-A-W/F-S-P) rich in aromatic amino acids. Four peptides, corresponding to the affinity selected sequences, were chemically synthesized and characterized for their carbohydrate recognition properties. The synthetic peptides exhibited high specificities and affinities to T antigen displayed on asialofetuin or conjugated to bovine serum albumin (Kd = 5 nM for MAP-P30 binding to asialofetuin) as well as free T-antigen disaccharide in solution (Kd = 10 microM for MAP-P30, 20 microM for P10). Two peptides, P30 and P10, demonstrated high affinities and specificities for both asialofetuin and T antigen in solution. Iodination of a lone tyrosine residue in each sequence dramatically reduced their abilities to bind T antigen, suggesting that the tyrosine residue plays an important role in carbohydrate recognition. That these peptides are of functional significance is evidenced by the ability of both P30 and P10 to inhibit asialofetuin-mediated melanoma cell aggregation in vitro and to compete with peanut lectin for binding to T antigen displayed on the surface of MDA-MB-435 breast carcinoma cells in situ.