Zuckermann R N, Kerr J M, Siani M A, Banville S C, Santi D V
Proc Natl Acad Sci U S A. 1992 May 15;89(10):4505-9. doi: 10.1073/pnas.89.10.4505.
A fully automated peptide synthesizer has been constructed that is capable of the simultaneous synthesis of up to 36 individual peptides and the synthesis of equimolar peptide mixtures. The instrument consists of an array of reaction vessels, a series of solenoid valves to control liquid flow, and a Zymark robot to deliver solvents and reagents; all components are computer controlled and coordinated. Equimolar peptide mixtures are obtained by algorithms that automate the mixing and distribution of peptide-resin particles. This technology was used to synthesize a library of 361 peptides, generated by randomizing two critical binding residues of a 10-mer epitope known to bind an anti-human immuno-deficiency virus gp120 monoclonal antibody. Each critical residue was substituted with 19 amino acids consisting of all the natural amino acids except cysteine. The library was synthesized as 19 pools, each containing 19 peptides. Each pool was screened in a solution-phase competition ELISA assay. The 12 most inhibitory peptides in the library were isolated by a rapid affinity-selection method and were identified by mass spectrometry and amino acid analysis. The binding properties of these 12 selected peptides were verified by synthesis and assay of the individual peptides. The two critical residues investigated were found to contribute independently to antibody binding.
已构建了一台全自动肽合成仪，它能够同时合成多达36种单独的肽以及等摩尔肽混合物。该仪器由一系列反应容器、用于控制液体流动的一系列电磁阀以及用于输送溶剂和试剂的Zymark机器人组成；所有组件均由计算机控制和协调。通过使肽 - 树脂颗粒的混合和分配自动化的算法获得等摩尔肽混合物。这项技术用于合成一个包含361种肽的文库，该文库通过随机化已知与抗人免疫缺陷病毒gp120单克隆抗体结合的10聚体表位的两个关键结合残基而产生。每个关键残基被19种氨基酸取代，这19种氨基酸由除半胱氨酸外的所有天然氨基酸组成。该文库被合成为19个池，每个池包含19种肽。每个池在溶液相竞争ELISA试验中进行筛选。通过快速亲和选择方法分离文库中12种抑制性最强的肽，并通过质谱和氨基酸分析进行鉴定。通过合成和分析单个肽验证了这12种选定肽的结合特性。发现所研究的两个关键残基对抗体结合有独立贡献。
