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Specificity of Subtilin-Mediated Activation of Histidine Kinase SpaK.

ABSTRACT: Autoinduction via two-component systems is a widespread regulatory mechanism that senses environmental and metabolic changes. Although the lantibiotics nisin and subtilin are closely related and share the same lanthionine ring structure, they autoinduce their biosynthesis in a highly specific manner. Subtilin activates only the two-component system SpaRK of Bacillus subtilis, whereas nisin activates solely the two-component system NisRK of Lactococcus lactis To identify components that determine the specificity of subtilin autoinduction, several variants of the respective lantibiotics were analyzed for their autoinductive capacities. Here, we show that amino acid position 20 is crucial for SpaK activation, as an engineered nisin molecule with phenylalanine at position 20 (nisin N20F) was able to activate SpaK in a specific manner. In combination with the N-terminal tryptophan of subtilin (nisin I1W/N20F), SpaK autoinduction reached almost the level of subtilin-mediated autoinduction. Furthermore, the overall structure of subtilin is also important for its association with the histidine kinase. The destruction of the second lanthionine ring (subtilin C11A, ring B), as well as mutations that interfere with the flexibility of the hinge region located between lanthionine rings C and D (subtilin L21P/Q22P), abolished SpaK autoinduction. Although the C-terminal part of subtilin is needed for efficient SpaK autoinduction, the destruction of lanthionine rings D and E had no measurable impact. Based on these findings, a model for the interaction of subtilin with histidine kinase SpaK was established.IMPORTANCE Although two-component systems are important regulatory systems that sense environmental changes, very little information on the molecular mechanism of sensing or the interaction of the sensor with its respective kinase is available. The strong specificity of linear lantibiotics such as subtilin and nisin for their respective kinases provides an excellent model system to unravel the structural needs of these lantibiotics for activating histidine kinases in a specific manner. More than that, the biosyntheses of lantibiotics are autoinduced via two-component systems. Therefore, an understanding of their interactions with histidine kinases is needed for the biosynthesis of newly engineered peptide antibiotics. Using a Bacillus subtilis-based reporter system, we were able to identify the molecular constraints that are necessary for specific SpaK activation and to provide SpaK specificity to nisin with just two point mutations.

SUBMITTER: Geiger C

PROVIDER: S-EPMC5583494 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Specificity of Subtilin-Mediated Activation of Histidine Kinase SpaK.

Geiger Christoph C Spieß Tobias T Korn Sophie Marianne SM Kötter Peter P Entian Karl-Dieter KD

Applied and environmental microbiology 20170831 18

Autoinduction via two-component systems is a widespread regulatory mechanism that senses environmental and metabolic changes. Although the lantibiotics nisin and subtilin are closely related and share the same lanthionine ring structure, they autoinduce their biosynthesis in a highly specific manner. Subtilin activates only the two-component system SpaRK of <i>Bacillus subtilis</i>, whereas nisin activates solely the two-component system NisRK of <i>Lactococcus lactis</i> To identify components ...[more]

PMID: 28710266

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Project description:BackgroundProtein kinase D (PKD) constitutes a novel family of serine/threonine protein kinases implicated in fundamental biological activities including cell proliferation, survival, migration, and immune responses. Activation of PKD in these cellular activities has been linked to many extracellular signals acting through antigen receptor engagement, receptor tyrosine kinases, as well as G protein-coupled receptors. In the latter case, it is generally believed that the G? subunits of the Gq family are highly effective in mediating PKD activation, whereas little is known with regard to the ability of G?? dimers and other G? subunits to stimulate PKD. It has been suggested that the interaction between G?? and the PH domain of PKD, or the G??-induced PLC?/PKC activity is critical for the induction of PKD activation. However, the relative contribution of these two apparently independent events to G??-mediated PKD activation has yet to be addressed.ResultsIn this report, we demonstrate that among various members in the four G protein families, only the G? subunits of the Gq family effectively activate all the three PKD isoforms (PKD1/2/3), while G? subunits of other G protein families (Gs, Gi, and G12) are ineffective. Though the G? subunits of Gi family are unable to stimulate PKD, receptors linked to Gi proteins are capable of triggering PKD activation in cell lines endogenously expressing (HeLa cells and Jurkat T-cells) or exogenously transfected with (HEK293 cells) G??-sensitive PLC?2/3 isoforms. This indicates that the Gi-mediated PKD activation is dependent on the released G?? dimers upon stimulation. Further investigation on individual G?? combinations (i.e. G?1 with G?1-13) revealed that, even if they can stimulate the PLC? activity in a comparable manner, only those G?1? dimers with ?2, ?3, ?4, ?5, ?7, and ?10 can serve as effective activators of PKD. We also demonstrated that Gi-mediated PKD activation is essential for the SDF-1?-induced chemotaxis on Jurkat T-cells.ConclusionsOur current report illustrates that G?? dimers from the Gi proteins may activate PKD in a PLC?2/3-dependent manner, and the specific identities of G? components within G?? dimers may determine this stimulatory action.

Dataset Information

Specificity of Subtilin-Mediated Activation of Histidine Kinase SpaK.

Publications

Specificity of Subtilin-Mediated Activation of Histidine Kinase SpaK.

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