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Efficient recombinant expression of SFTI ‐1 in bacterial cells using intein‐mediated protein trans‐splicing
Author(s) -
Li Yilong,
Aboye Teshome,
Breindel Leonard,
Shekhtman Alexander,
Camarero Julio A.
Publication year - 2016
Publication title -
peptide science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/bip.22875
Subject(s) - intein , cyclic peptide , chemistry , protein splicing , intracellular , recombinant dna , peptide , biochemistry , rna splicing , directed evolution , protein tag , trypsin , microbiology and biotechnology , biology , gene , mutant , enzyme , fusion protein , rna
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 µ M . 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.