Martín María, Gutiérrez Jorge, Criado Raquel, Herranz Carmen, Cintas Luis M, Hernández Pablo E
Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Complutense de Madrid, 28040, Madrid, Spain.
Appl Microbiol Biotechnol. 2007 Sep;76(3):667-75. doi: 10.1007/s00253-007-1044-3. Epub 2007 Jun 27.
Replacement of the leader sequence of enterocin A (EntA), a bacteriocin produced by Enterococcus faecium PLBC21, by the signal peptide of enterocin P (EntP), a sec-dependent bacteriocin produced by E. faecium P13, permitted production of EntA in Lactococcus lactis. Chimeras encoding the EntP signal peptide (SP( entP )) fused to mature EntA (entA), with or without its immunity gene (entiA), were cloned into the expression vector pMG36c to generate the recombinant plasmids, pMPA15 (SP( entP ):entA) and pMPA10i (SP( entP ):entA + entiA). Transformation of competent L. lactis subsp. lactis IL1403 and L. lactis subsp. cremoris NZ9000 with the recombinant plasmids permitted production of EntA by the transformed cells, and the co-production of nisin A and EntA by the L. lactis subsp. lactis DPC5598 transformants. Mature EntA fused to SP(EntP) is the minimum requirement for synthesis, processing and secretion of biologically active EntA in L. lactis. The production of EntA by most recombinant L. lactis hosts was larger than in the E. faecium control strains. All L. lactis derivatives showed antimicrobial activity against Listeria spp., and L. lactis (pMPA15) displayed the highest antilisterial effect.
用粪肠球菌P13产生的一种依赖Sec的细菌素肠球菌素P（EntP）的信号肽替换粪肠球菌PLBC21产生的细菌素肠球菌素A（EntA）的前导序列，使得乳酸乳球菌能够产生EntA。将编码与成熟EntA（entA）融合的EntP信号肽（SP(entP)）的嵌合体，无论有无其免疫基因（entiA），克隆到表达载体pMG36c中，以产生重组质粒pMPA15（SP(entP):entA）和pMPA10i（SP(entP):entA + entiA）。用重组质粒转化乳酸乳球菌乳酸亚种IL1403和乳酸乳球菌乳脂亚种NZ9000，使得转化细胞能够产生EntA，并且乳酸乳球菌乳酸亚种DPC5598转化子能够共同产生乳酸链球菌素A和EntA。与SP(EntP)融合的成熟EntA是在乳酸乳球菌中合成、加工和分泌具有生物活性的EntA的最低要求。大多数重组乳酸乳球菌宿主产生的EntA比粪肠球菌对照菌株中的产量高。所有乳酸乳球菌衍生物均对李斯特菌属表现出抗菌活性，并且乳酸乳球菌（pMPA15）表现出最高的抗李斯特菌效果。
