In vivo biosynthesis of an Ala-scan library based on the cyclic peptide SFTI-1.
We present the in vivo biosynthesis of wild-type sunflower trypsin inhibitor 1 (SFTI-1) inside E. coli cells using an intramolecular native chemical ligation in combination with a modified protein splicing unit. SFTI-1 is a small backbone cyclized polypeptide with a single disulfide bridge. A small library containing multiple Ala mutants was also biosynthesized and its activity was assayed using a trypsin-binding assay. This study clearly demonstrates the exciting possibility of generating large cyclic peptide libraries in live E. coli cells, and is a critical first step for developing in vivo screening and directed evolution technologies using the cyclic peptide SFTI-1 as a molecular scaffold.
Austin
Jeffrey
J
Lawrence Livermore National Laboratory, Livermore, CA 94550, USA.
Kimura
Richard H
RH
Woo
Youn-Hi
YH
Camarero
Julio A
JA
D000409:Alanine / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D017931:DNA Primers; D008969:Molecular Sequence Data; D019151:Peptide Library; D010456:Peptides, Cyclic / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization
D017931:DNA Primers; D019151:Peptide Library; D010456:Peptides, Cyclic; C448501:SFTI-1 peptide, sunflower; D000409:Alanine
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