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Analysis of the Bacillus cereus SpoIIS antitoxin-toxin system reveals its three-component nature.

Melničáková J, Bečárová Z, Makroczyová J, Barák I - Front Microbiol (2015)

Bottom Line: In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC.Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system.In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Slovak Academy of Sciences Bratislava, Slovakia.

ABSTRACT
Programmed cell death in bacteria is generally associated with two-component toxin-antitoxin systems. The SpoIIS toxin-antitoxin system, consisting of a membrane-bound SpoIISA toxin and a small, cytosolic antitoxin SpoIISB, was originally identified in Bacillus subtilis. In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC. Its protein product serves as an antitoxin, and similarly as SpoIISB, is able to bind SpoIISA and abolish its toxic effect. Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system. In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

No MeSH data available.


Related in: MedlinePlus

Analysis of the multimeric state of B. cereus C-SpoIISA. (A) Dynamic light scattering analysis of C-SpoIISA oligomer. Size distribution (by intensity) of B. cereus C-SpoIISA, at 20°C, average hydrodynamic radius = 55 nm. (B) Gel filtration of C-SpoIISA. The stars indicate the positions at which the following protein standards eluted from the column (left to right): 2000, 450, 66, 45, and 29 kDa. (C) Western blot analysis of glutaraldehyde-crosslinked His6-tagged C-SpoIISA.
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Figure 5: Analysis of the multimeric state of B. cereus C-SpoIISA. (A) Dynamic light scattering analysis of C-SpoIISA oligomer. Size distribution (by intensity) of B. cereus C-SpoIISA, at 20°C, average hydrodynamic radius = 55 nm. (B) Gel filtration of C-SpoIISA. The stars indicate the positions at which the following protein standards eluted from the column (left to right): 2000, 450, 66, 45, and 29 kDa. (C) Western blot analysis of glutaraldehyde-crosslinked His6-tagged C-SpoIISA.

Mentions: The crystal structure of the B. subtilis SpoIISA C-terminal domain shows that the protein dimerizes by forming a four-helix bundle using the first and last α-helices of each molecule (Florek et al., 2011). Our bacterial two-hybrid experiments showed that B. cereus C-SpoIISA interacts with other B. cereus C-SpoIISA molecules (Figure 3), suggesting that this molecule also forms oligomers. The oligomeric form of C-SpoIISA was examined by measuring the hydrodynamic radius of dissolved particles using dynamic light scattering. A cumulant analysis showed that the sample was monomodal (i.e., had only one peak, Figure 5A), and was polydisperse, with a polydispersity index of 0.255 and an overall polydispersity of 50.32%. The polydispersity indicates broader particle size distribution, and thus the hydrodynamic radius and corresponding molecular mass cannot be reliably calculated.


Analysis of the Bacillus cereus SpoIIS antitoxin-toxin system reveals its three-component nature.

Melničáková J, Bečárová Z, Makroczyová J, Barák I - Front Microbiol (2015)

Analysis of the multimeric state of B. cereus C-SpoIISA. (A) Dynamic light scattering analysis of C-SpoIISA oligomer. Size distribution (by intensity) of B. cereus C-SpoIISA, at 20°C, average hydrodynamic radius = 55 nm. (B) Gel filtration of C-SpoIISA. The stars indicate the positions at which the following protein standards eluted from the column (left to right): 2000, 450, 66, 45, and 29 kDa. (C) Western blot analysis of glutaraldehyde-crosslinked His6-tagged C-SpoIISA.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526809&req=5

Figure 5: Analysis of the multimeric state of B. cereus C-SpoIISA. (A) Dynamic light scattering analysis of C-SpoIISA oligomer. Size distribution (by intensity) of B. cereus C-SpoIISA, at 20°C, average hydrodynamic radius = 55 nm. (B) Gel filtration of C-SpoIISA. The stars indicate the positions at which the following protein standards eluted from the column (left to right): 2000, 450, 66, 45, and 29 kDa. (C) Western blot analysis of glutaraldehyde-crosslinked His6-tagged C-SpoIISA.
Mentions: The crystal structure of the B. subtilis SpoIISA C-terminal domain shows that the protein dimerizes by forming a four-helix bundle using the first and last α-helices of each molecule (Florek et al., 2011). Our bacterial two-hybrid experiments showed that B. cereus C-SpoIISA interacts with other B. cereus C-SpoIISA molecules (Figure 3), suggesting that this molecule also forms oligomers. The oligomeric form of C-SpoIISA was examined by measuring the hydrodynamic radius of dissolved particles using dynamic light scattering. A cumulant analysis showed that the sample was monomodal (i.e., had only one peak, Figure 5A), and was polydisperse, with a polydispersity index of 0.255 and an overall polydispersity of 50.32%. The polydispersity indicates broader particle size distribution, and thus the hydrodynamic radius and corresponding molecular mass cannot be reliably calculated.

Bottom Line: In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC.Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system.In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Slovak Academy of Sciences Bratislava, Slovakia.

ABSTRACT
Programmed cell death in bacteria is generally associated with two-component toxin-antitoxin systems. The SpoIIS toxin-antitoxin system, consisting of a membrane-bound SpoIISA toxin and a small, cytosolic antitoxin SpoIISB, was originally identified in Bacillus subtilis. In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC. Its protein product serves as an antitoxin, and similarly as SpoIISB, is able to bind SpoIISA and abolish its toxic effect. Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system. In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

No MeSH data available.


Related in: MedlinePlus