Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, P.O. Box 5003, 1432, Ås, Norway; Molecular Genetics Group, Groningen Biomolecular Sciences and Biotechnology Institute, Centre for Synthetic Biology, University of Groningen, Nijenborgh 7, 9747 AG, Groningen, The Netherlands.
Mol Microbiol. 2014 Jun;92(6):1177-87. doi: 10.1111/mmi.12632. Epub 2014 May 23.
Most bacterially produced antimicrobial peptides (bacteriocins) are thought to kill target cells by a receptor-mediated mechanism. However, for most bacteriocins the receptor is unknown. For instance, no target receptor has been identified for the two-peptide bacteriocins (class IIb), whose activity requires the combined action of two individual peptides. To identify the receptor for the class IIb bacteriocin lactococcin G, which targets strains of Lactococcus lactis, we generated 12 lactococcin G-resistant mutants and performed whole-genome sequencing to identify mutations causing the resistant phenotype. Remarkably, all had a mutation in or near the gene uppP (bacA), encoding an undecaprenyl pyrophosphate phosphatase; a membrane protein involved in peptidoglycan synthesis. Nine mutants had stop codons or frameshifts in the uppP gene, two had point mutations in putative regulatory regions and one caused an amino acid substitution in UppP. To verify the receptor function of UppP, it was shown that growth of non-sensitive Streptococcus pneumoniae could be inhibited by lactococcin G when L. lactis uppP was expressed in this bacterium. Furthermore, we show that the related class IIb bacteriocin enterocin 1071 also uses UppP as receptor. The approach used here should be broadly applicable to identify receptors for other bacteriocins as well.
大多数由细菌产生的抗菌肽（细菌素）被认为通过受体介导的机制杀死靶细胞。然而，对于大多数细菌素来说，受体是未知的。例如，尚未鉴定出针对双肽细菌素（IIb 类）的靶受体，其活性需要两个单独肽的共同作用。为了鉴定靶向乳球菌（Lactococcus lactis）菌株的 IIb 类细菌素乳球菌素 G 的受体，我们生成了 12 个乳球菌素 G 抗性突变体，并进行全基因组测序以鉴定导致抗性表型的突变。值得注意的是，所有突变体在基因 uppP（bacA）中或附近都有突变，该基因编码一种十一碳烯焦磷酸磷酸酶；一种参与肽聚糖合成的膜蛋白。9 个突变体在 uppP 基因中具有终止密码子或移码突变，2 个突变体在假定的调节区域中有点突变，1 个突变体导致 UppP 中的氨基酸取代。为了验证 UppP 的受体功能，表明当在这种细菌中表达乳球菌的 lactisuppP 时，非敏感肺炎链球菌的生长可以被乳球菌素 G 抑制。此外，我们表明相关的 IIb 类细菌素肠菌素 1071 也使用 UppP 作为受体。此处使用的方法应该广泛适用于鉴定其他细菌素的受体。
