Title:Streptomyces as Overexpression System for Heterologous Production of an Antimicrobial Peptide
Volume: 24
Issue: 6
Author(s): Marisol Roldán-Tapia, Jozef Anné, Ana Gisela Reyes, Ulises Carrasco, Cesar Millán-Pacheco, Javier Barrios-González and Armando Mejía*
Affiliation:
- Department of Biotechnology, Universidad Autónoma Metropolitana San Rafael Atlixco 186, Vicentina, Iztapalapa, P.O. Box: 09340. Ciudad de México,Mexico
Keywords:
Heterologous production, antimicrobial peptide, Streptomyces lividans, secreted peptide, phytopathogens, CAP
overexpression.
Abstract: Background: Antimicrobial peptides could be used in several fields of application, and
large quantities of antimicrobial peptides would be required. However, their production is very expensive;
this is why a suitable production method, alternative to traditional chemical synthesis is
necessary. Production of recombinant antimicrobial peptides in prokaryotic systems has demonstrated
the viability of this approach. Nevertheless, expression of antimicrobial peptides in Escherichia
coli an others microorganisms is potentially limited due to their toxicity to host cells and
susceptibility to proteolytic degradation. As an alternative, we describe a successful antimicrobial
peptide production system in Streptomyces lividans which showed to be effective for the secretion of
large quantities of cationic antimicrobial peptides.
Objective: Therefore, as a solution to the difficulties for heterologous expression of CAP we demonstrate
efficient production by
S. lividans.
Method: In this study, a strategy for CAP overexpression is presented based on the construction of
an expression cassette for
Streptomyces lividans TK24. For the construction of this cassette, the peptide
of interest was fused to the vsi promoter and signal sequence
(vsi-ss) of the subtilisin inhibitor
from
Streptomyces venezuelae CBS762.70, which is a signal peptide with a proven high secretion
efficiency. The cloning vector used was pIJ486, which includes a transcription terminator sequence
and a thiostrepton resistance marker. This system contains elements that allow the increase of the
efficiency of the peptide’s expression.
Results: The production system allows the efficient secretion of the peptide to the growth medium,
thereby simplifying its recovery and avoiding its toxic effect on the producing organism. The production
obtained demonstrated the system’s efficiency by achieving a peptide concentration of 11.61
mg/ml. This represents at least a 10-fold increase compared to previously established strategies.
Conclusion: The expression system constructed may facilitate the production of large amounts of
peptides with antimicrobial activity.
