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Chemical and Biosynthetic Approaches to the Production of Novel Polypeptide Materials
Author(s) -
Mcgrath Kevin P.,
Tirrell David A.,
Kawai Michio,
Mason Thomas L.,
Fournier Maurille J.
Publication year - 1990
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp00003a004
Subject(s) - plasmid , sequence (biology) , recombinant dna , chemical synthesis , dna , transformation (genetics) , insert (composites) , combinatorial chemistry , population , chemistry , biology , stereochemistry , biochemistry , gene , materials science , demography , sociology , in vitro , composite material
Abstract Three approaches to the synthesis of the repetitive copolypeptide [‐(GlyAla) 3‐ GlyProGlu‐] n (1) are described. Direct chemical synthesis of 1 via classical solution methods required 18 steps and afforded a polydisperse product with an average molecular weight of less than 10 000. Two alternative genetic strategies were also explored. In the first, chemically synthesized DNA oligomers were self‐ligated to produce a population of multimers, which were fitted with translational start and stop signals and inserted into an expression plasmid containing the λ P L promoter and a synthetic ribosome binding site. Transformation of E. coli led to the isolation of a stable recombinant plasmid carrying an insert encoding 12 repeats of sequence 1. Attempts to identify polypeptide 1 after induction of transformed cultures were unsuccessful. A second strategy, generating a tripartite derivative of sequence 1 carrying short N‐and C‐terminal extensions, afforded excellent yields of product. The relative merits of chemical and genetic approaches to repetitive polypeptide materials are discussed.