title: Efficient recombinant expression of SFTI-1 in bacterial cells using intein-mediated protein trans-splicing.
abstract: We report for the first time the recombinant expression of bioactive wild-type sunflower trypsin inhibitor 1 (SFTI-1) inside E. coli cells by making use of intracellular protein trans-splicing in combination with a high efficient split-intein. SFTI-1 is a small backbone-cyclized polypeptide with a single disulfide bridge and potent trypsin inhibitory activity. Recombinantly produced SFTI-1 was fully characterized by NMR and was observed to actively inhibit trypsin. The in-cell expression of SFTI-1 was very efficient reaching intracellular concentration ≈ 40 µM. This study clearly demonstrates the possibility of generating genetically encoded SFTI-based peptide libraries in live E. coli cells, and is a critical first step for developing in-cell screening and directed evolution technologies using the cyclic peptide SFTI-1 as a molecular scaffold. © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 818-824, 2016.
authors.affiliation: Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, CA, 90089-9121.
authors.forename: Yilong
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, CA, 90089-9121.
authors.forename: Teshome
authors.initials: T
authors.lastname: Aboye
authors.affiliation: Department of Chemistry, State University of New York, Albany, NY, 12222.
authors.forename: Leonard
authors.initials: L
authors.lastname: Breindel
authors.affiliation: Department of Chemistry, State University of New York, Albany, NY, 12222.
authors.forename: Alexander
authors.initials: A
authors.lastname: Shekhtman
authors.affiliation: Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, CA, 90089-9121.|Department of Chemistry, University of Southern California, Los Angeles, CA, 90089-9121.
authors.forename: Julio A
authors.initials: JA
authors.lastname: Camarero
mesh_terms: D004926:Escherichia coli; D015870:Gene Expression; D006368:Helianthus / Q000737:chemistry / Q000235:genetics; D047668:Inteins; D010456:Peptides, Cyclic / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D019154:Protein Splicing; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics
chemical_list: D010456:Peptides, Cyclic; D011994:Recombinant Proteins; C448501:SFTI-1 peptide, sunflower
keywords: Bowman-Birk inhibitor; backbone-cyclized peptides; protein splicing; split-intein; sunflower trypsin inhibitor