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Co-expression and characterization of enterocin CRL35 and its mutant in Escherichia coli Rosetta

Emilse Masías
  • Instituto Superior de Investigaciones Biologicas (CONICET-UNT) e Instituto de Quimica Biologica ‘‘Dr. Bernabe Bloj’’, San Miguel de Tucuman, Argentina
  • Other articles by this author:
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/ Gianluca Picariello
  • Istituto di Scienze dell’Alimentazione – Consiglio Nazionale delle Ricerche (CNR), Via Roma, 64 – 83100 Avellino, Italy
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  • De Gruyter OnlineGoogle Scholar
/ Leonardo Acuña
  • Instituto Superior de Investigaciones Biologicas (CONICET-UNT) e Instituto de Quimica Biologica ‘‘Dr. Bernabe Bloj’’, San Miguel de Tucuman, Argentina
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  • De Gruyter OnlineGoogle Scholar
/ Miriam Chalón
  • Instituto Superior de Investigaciones Biologicas (CONICET-UNT) e Instituto de Quimica Biologica ‘‘Dr. Bernabe Bloj’’, San Miguel de Tucuman, Argentina
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  • De Gruyter OnlineGoogle Scholar
/ Fernando Sesma / Roberto Morero
  • Instituto Superior de Investigaciones Biologicas (CONICET-UNT) e Instituto de Quimica Biologica ‘‘Dr. Bernabe Bloj’’, San Miguel de Tucuman, Argentina
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/ Lucila Saavedra / Carlos Minahk
  • Corresponding author
  • Chacabuco 461, S.M. de Tucuman (T4000ILI), Argentina, Fax: +54 0381 4248921, Tel: +54 0381 4248921
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Published Online: 2014-04-24 | DOI: https://doi.org/10.2478/ped-2014-0003


Even though many sequences and structures of bacteriocins from lactic acid bacteria have been fully characterized so far, little information is currently available about bacteriocins heterologously produced by Escherichia coli. For this purpose, the structural gene of enterocin CRL35, munA, was PCR-amplified using specific primers and cloned downstream of PelB sequence in the pET22b (+) expression vector. E. coli Rosetta (DE3) pLysS was chosen as the host for production and enterocin was purified by an easy two-step protocol. The bacteriocin was correctly expressed with the expected intramolecular disulfide bond. Nevertheless, it was found that a variant of the enterocin, differing by 12 Da from the native polypeptide, was co-expressed by E. coli Rosetta in comparable amount. Indeed, the mutant bacteriocin contained two amino acid substitutions that were characterized by matrix assisted laser desorption ionization-time of flight (MALDI-TOF) and HPLCelectrospray (ESI)-Q-TOF tandem mass spectrometry (MS/ MS) sequencing. This is the first report regarding the production of mutants of pediocin-like bacteriocins in the E. coli expression system.

Keywords : bacteriocins; heterologous expression; MALDITOF MS analysis; nanoHPLC-ESI MS/MS analysis


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About the article

Received: 2013-07-26

Accepted: 2013-12-06

Published Online: 2014-04-24

Published in Print: 2014-01-01

Citation Information: Peptidomics, Volume 1, Issue 1, ISSN (Online) 2084-7203, DOI: https://doi.org/10.2478/ped-2014-0003.

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© 2014 Emilse Masías et al.. This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License. BY-NC-ND 3.0

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