title: In vivo biosynthesis of an Ala-scan library based on the cyclic peptide SFTI-1.
abstract: 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.
authors.affiliation: Lawrence Livermore National Laboratory, Livermore, CA 94550, USA.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Austin
authors.forename: Richard H
authors.initials: RH
authors.lastname: Kimura
authors.forename: Youn-Hi
authors.initials: YH
authors.lastname: Woo
authors.forename: Julio A
authors.initials: JA
authors.lastname: Camarero
mesh_terms: 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
chemical_list: D017931:DNA Primers; D019151:Peptide Library; D010456:Peptides, Cyclic; C448501:SFTI-1 peptide, sunflower; D000409:Alanine
references.citation: J Mol Biol. 1999 Dec 17;294(5):1327-36
references.pmid: 10600388
references.citation: J Biochem. 2000 Dec;128(6):1051-7
references.pmid: 11098149
references.citation: J Bacteriol. 2003 Jul;185(14):4011-21
references.pmid: 12837774
references.citation: Proc Natl Acad Sci U S A. 1999 Nov 23;96(24):13638-43
references.pmid: 10570125
references.citation: Biochem Pharmacol. 1973 Dec 1;22(23):3099-108
references.pmid: 4202581
references.citation: J Biol Chem. 2003 Jun 13;278(24):21782-9
references.pmid: 12621047
references.citation: Biochemistry. 2005 Feb 1;44(4):1145-53
references.pmid: 15667208
references.citation: Curr Protein Pept Sci. 2004 Oct;5(5):351-64
references.pmid: 15544530
references.citation: Angew Chem Int Ed Engl. 2006 Jan 30;45(6):973-6
references.pmid: 16381044
references.citation: Proc Natl Acad Sci U S A. 1998 Aug 4;95(16):9226-31
references.pmid: 9689062
references.citation: Trends Biochem Sci. 2002 Mar;27(3):132-8
references.pmid: 11893510
references.citation: Annu Rev Biochem. 1992;61:387-418
references.pmid: 1497316
references.citation: Bioorg Med Chem. 2001 Sep;9(9):2479-84
references.pmid: 11553489
references.citation: Protein Sci. 1998 Nov;7(11):2256-64
references.pmid: 9827992
references.citation: J Biol Chem. 1998 Jun 26;273(26):16205-9
references.pmid: 9632677
references.citation: Proc Natl Acad Sci U S A. 1998 Jun 9;95(12):6705-10
references.pmid: 9618476
references.citation: Curr Protoc Protein Sci. 2001 May;Chapter 18:Unit18.4
references.pmid: 18429140
references.citation: Angew Chem Int Ed Engl. 1998 Feb 16;37(3):347-349
references.pmid: 29711251
references.citation: FEBS Lett. 1999 Oct 8;459(2):166-72
references.pmid: 10518012
references.citation: Curr Opin Drug Discov Devel. 2006 Mar;9(2):251-60
references.pmid: 16566295
references.citation: J Pept Res. 1998 Apr;51(4):303-16
references.pmid: 9560006
references.citation: J Mol Biol. 1999 Jul 9;290(2):525-33
references.pmid: 10390350
references.citation: Curr Opin Drug Discov Devel. 2002 Mar;5(2):251-60
references.pmid: 11926131
references.citation: Chembiochem. 2007 Aug 13;8(12):1363-6
references.pmid: 17590879
references.citation: Curr Protein Pept Sci. 2005 Feb;6(1):7-21
references.pmid: 15638765
references.citation: Protein Eng Des Sel. 2004 Sep;17(9):681-7
references.pmid: 15486024
references.citation: J Biol Chem. 2000 Mar 31;275(13):9091-4
references.pmid: 10734038
references.citation: J Mol Biol. 2001 Aug 17;311(3):579-91
references.pmid: 11493011
references.citation: Biochem J. 1990 Jun 1;268(2):249-62
references.pmid: 2163604
references.citation: Anal Biochem. 2007 Oct 1;369(1):60-70
references.pmid: 17586456
references.citation: Biopolymers. 2006;84(3):250-66
references.pmid: 16440288
references.citation: J Biol Chem. 2006 Aug 18;281(33):23668-75
references.pmid: 16766795
references.citation: Chembiochem. 2002 Apr 2;3(4):318-23
references.pmid: 11933232