Expression of five class II bacteriocins with activity against Escherichia coli in Lacticaseibacillus paracasei CNCM I-5369, and in a heterologous host

Abstract

Highlights Lacticaseibacillus paracasei CNCM I-5369 produces five class II bacteriocins. Genes coding these class II bacteriocins are expressed in the natural host. These bacteriocins are individually produced in E. coli as heterologous host. These class II bacteriocins are active against E. coli . Together, these class II bacteriocins are synergistic.

Lacticaseicin 30 and Colistin as a Promising Antibiotic Formulation against Gram-Negative β-Lactamase-Producing Strains and Colistin-Resistant Strains

“…E. coli Rosetta -T7-6his-30 strain ( Table 3 ) was used to produce recombinant lacticaseicin 30. This strain was grown in Luria-Bertani broth (LB) at 37 °C with shaking at 160 rpm [ 25 ].…”

“…To recover the soluble recombinant bacteriocin fraction from the cytoplasm, cells were lysed by sonication three times during 40 s at 180 Watt (OmniRuptor 4000 Ultrasonic Homogenizer, Omni International, Georgia, GA, USA). After their lysis, the bacteriocin was purified by the Ni-NTA chromatography as recently reported by Madi-Moussa et al [ 25 ]. The histidine tag was removed with Tev protease (Sigma-Aldrich) following the recommended instructions.…”

“…The Ct value is the basis for the calculation of the relative quantification, corresponding to the expression of the target gene ( mcr gene) compared with the house-keeping gene ( 16S rRNA gene), as demonstrated in [ 28 ]. The analysis of the relative expression of the target genes was determined using the 2 −(ΔΔCt) method as previously described by [ 25 , 80 ].…”

“…Bacteriocins produced by L. paracasei CNCM I-5369 were successfully produced in a heterologous host Escherichia coli Rosetta by Madi-Moussa [ 25 ]. Bacteriocin coded by orf30 was particularly important because of its easy production in large quantities, unlike other ones encoded by orf010 , orf012 , orf023 and orf038 [ 25 ].…”

“…Bacteriocins produced by L. paracasei CNCM I-5369 were successfully produced in a heterologous host Escherichia coli Rosetta by Madi-Moussa [ 25 ]. Bacteriocin coded by orf30 was particularly important because of its easy production in large quantities, unlike other ones encoded by orf010 , orf012 , orf023 and orf038 [ 25 ]. Noteworthy, bacteriocin 30, hence designed as lacticaseicin 30, was assessed in this work alone and in conjunction with colistin against a set of Gram-negative pathogenic bacteria from a clinical source, in order to design a potential therapeutic issue.…”