Amino Acids

, Volume 38, Issue 5, pp 1313–1322 | Cite as

In vivo biosynthesis of an Ala-scan library based on the cyclic peptide SFTI-1

  • Jeffrey Austin
  • Richard H. Kimura
  • Youn-Hi Woo
  • Julio A. Camarero
Original Article


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.


Bowman–Birk inhibitor Trypsin inhibitor Backbone cyclized peptides Genetically encoded libraries Protein splicing 





Chitin-binding domain




Cyclic cystine-knot motif


Bowman–Birk inhibitor


Ethylenediaminetetraacetic acid




Guanidinium hydrochloride




High performance liquid chromatography




Methionyl aminopeptidase


Mercaptoethanesulfonic acid


N-hydroxysuccinimide ester


Polyacrylamide gel electrophoresis


Phenylmethylsulfonyl fluoride


Sodium dodecyl sulfate


Sunflower trypsin inhibitor 1


Trifluoroacetic acid





Work was supported by funding from the School of Pharmacy at the University of Southern California and Lawrence Livermore National Laboratory.

Supplementary material

726_2009_338_MOESM1_ESM.doc (587 kb)
Supplementary material 1 (DOC 587 kb)


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Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Jeffrey Austin
    • 2
  • Richard H. Kimura
    • 2
  • Youn-Hi Woo
    • 2
  • Julio A. Camarero
    • 1
    • 2
  1. 1.Department of Pharmaceutical Sciences and Pharmacology, School of PharmacyUniversity of Southern CaliforniaLos AngelesUSA
  2. 2.Lawrence Livermore National LaboratoryLivermoreUSA

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