DESIGN OF SYNTHETIC GENE LIBRARIES ENCODING RANDOM SEQUENCE PROTEINS WITH DESIRED ENSEMBLE CHARACTERISTICS
DESIGN OF SYNTHETIC GENE LIBRARIES ENCODING RANDOM SEQUENCE PROTEINS WITH DESIRED ENSEMBLE CHARACTERISTICS
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DOI：
10.1002/pro.5560020807
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发表时间：
1993-08-01
期刊：
影响因子：
8
通讯作者：
KAUFFMAN, SA
中科院分区：
文献类型：
--
作者：
LABEAN, TH;KAUFFMAN, SA
随机序列多肽文库可用作未进化蛋白质、新型配体和用于开发疫苗和治疗剂的潜在先导化合物的来源。之前已经使用用等摩尔核苷酸混合物合成的DNA实现了小随机肽的表达。对于随机多肽文库的许多潜在用途，诸如避免终止密码子和匹配靶氨基酸组成的问题使得需要更复杂的设计。在这项研究中，设计了三种核苷酸混合物，对应于密码子中的三个位置，使得通过三种混合物的重复循环合成的半随机DNA产生编码具有所需总体特征的随机序列多肽的开放阅读框架。使用两种方法来设计核苷酸混合物：手动使用电子表格和细化网格搜索算法。使用小于或等于1%终止密码子的设计目标和基于在天然球状蛋白中观察到的平均比率的氨基酸组成，搜索方法产生相似的核苷酸比率。半随机DNA，合成与设计，三个残基重复模式，可以编码非常高的多样性的图书馆，并代表了一个重要的工具，用于构建随机多肽文库。
Libraries of random sequence polypeptides are useful as sources of unevolved proteins, novel ligands, and potential lead compounds for the development of vaccines and therapeutics. The expression of small random peptides has been achieved previously using DNA synthesized with equimolar mixtures of nucleotides. For many potential uses of random polypeptide libraries, concerns such as avoiding termination codons and matching target amino acid compositions make more complex designs necessary. In this study, three mixtures of nucleotides, corresponding to the three positions in the codon, were designed such that semirandom DNA synthesized by repeated cycles of the three mixtures created an open reading frame encoding random sequence polypeptides with desired ensemble characteristics. Two methods were used to design the nucleotide mixtures: the manual use of a spreadsheet and a refining grid search algorithm. Using design targets of less than or equal to 1 % stop codons and an amino acid composition based on the average ratios observed in natural, globular proteins, the search methods yielded similar nucleotide ratios. Semirandom DNA, synthesized with a designed, three-residue repeat pattern, can encode libraries of very high diversity and represents an important tool for the construction of random polypeptide libraries.