Autoregulation of Lantibiotic Bovicin HJ50 Biosynthesis by the BovK-BovR Two-Component Signal Transduction System in Streptococcus bovis HJ50

Abstract

Streptococcus bovis HJ50 produces a lacticin 481-like 33-amino-acid-residue lantibiotic, designated bovicin HJ50. bovK-bovR in the bovicin HJ50 biosynthetic gene cluster is predicted to be a two-component signal transduction system involved in sensing signals and regulating gene expression. Disruption of bovK or bovR resulted in the abrogation of bovicin HJ50 production, suggesting both genes play important roles in bovicin HJ50 biosynthesis. Addition of exogenous bovicin HJ50 peptide to cultures of a bovM mutant that lost the capability for bovicin HJ50 production and structural gene bovA transcription in S. bovis HJ50 induced dose-dependent transcription of the bovA gene, demonstrating that bovicin HJ50 production was normally autoregulated. The transcription of bovA was no longer induced by bovicin HJ50 in bovK and bovR disruption mutants, suggesting that BovK-BovR plays an essential role in the signal transduction regulating bovicin HJ50 biosynthesis. A phosphorylation assay indicated that BovK has the ability to autophosphorylate and subsequently transfer the phosphoryl group to the downstream BovR protein to carry on signal transduction. Electromobility shift assays (EMSA) and green fluorescent protein (GFP) reporter gene expression assays showed the specific binding of BovR to the bovA promoter, indicating that BovR regulates bovA expression by direct binding between them. Taken together, bovicin HJ50 biosynthesis is induced by bovicin HJ50 itself and regulated via the two-component signal transduction system BovK-BovR.

Restoration of Bioactive Lantibiotic Suicin from a Remnant lan Locus of Pathogenic Streptococcus suis Serotype 2

“…5Ga and b). We speculate that it might be due to high-level expression of SuiK driven by strong lactococcal promoter P32, which leads to autophosphorylation of SuiK and signal leakage (24). Yet, SuiR alone was able to activate P suiA transcription, though at low efficiency, as was also observed with BovR and SpaR (24,48).…”

“…Like BovK-BovR, SalK in S. suis was presumed to be an eight-transmembrane kinase and SalR was a cytoplasmic regulator composed of an N-terminal signal receiver domain and a C-terminal HTH (helix-turn-helix) DNA binding domain. Both BovK-BovR of S. bovis and SalK-SalR of S. salivarius were demonstrated to be responsible for autoregulation of lantibiotic biosynthesis (24,31). Thus, we speculated that SalK-SalR in S. suis serotype 2 might be involved in lantibiotic production.…”

“…However, GFP expression was independent of suicin induction ( Fig. 5Ga and b), which might have been due to autophosphorylation of SuiK resulting from high-level expression of SuiK-SuiR driven by strong lactococcal promoter P32 (24). To see if SuiK was required for P suiA activation, pMG36e-suiRPsuiAgfp was constructed and transformed into L. lactis NZ9000, which resulted in much lower GFP expression (Fig.…”

“…SuiT 159 was N-terminally and SuiR was C-terminally 6ϫHis tagged. A suiA promoter (P suiA ) was amplified from genomic DNA of S. suis Habb and gfp from pGFP (24). P suiA and gfp were fused by overlapping PCR and introduced into pMG36e to obtain pMG36e-PsuiAgfp.…”

“…The assay was modified from an assay described previously (24). Briefly, pMG36e-derived plasmids containing suiKR or their mutants were transformed into L. lactis NZ9000 by the use of an electroporation method as described by Holo and Nes (30).…”