Structural basis of toxicity and immunity in contact-dependent growth inhibition (CDI) systems

Contact-dependent growth inhibition (CDI) systems encode polymorphic toxin/immunity proteins that mediate competition between neighboring bacterial cells. We present crystal structures of CDI toxin/immunity complexes from Escherichia coli EC869 and Burkholderia pseudomallei 1026b. Despite sharing little sequence identity, the toxin domains are structurally similar and have homology to endonucleases. The EC869 toxin is a Zn2+-dependent DNase capable of completely degrading the genomes of target cells, whereas the Bp1026b toxin cleaves the aminoacyl acceptor stems of tRNA molecules. Each immunity protein binds and inactivates its cognate toxin in a unique manner. The EC869 toxin/immunity complex is stabilized through an unusual β-augmentation interaction. In contrast, the Bp1026b immunity protein exploits shape and charge complementarity to occlude the toxin active site. These structures represent the initial glimpse into the CDI toxin/immunity network, illustrating how sequence-diverse toxins adopt convergent folds yet retain distinct binding interactions with cognate immunity proteins. Moreover, we present visual demonstration of CDI toxin delivery into a target cell.

[1]  Colin Kleanthous,et al.  Identification of the catalytic motif of the microbial ribosome inactivating cytotoxin colicin E3 , 2004, Protein science : a publication of the Protein Society.

[2]  C. Hayes,et al.  Identification of a target cell permissive factor required for contact-dependent growth inhibition (CDI). , 2012, Genes & development.

[3]  G. Moore,et al.  Cell entry mechanism of enzymatic bacterial colicins: porin recruitment and the thermodynamics of receptor binding. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[4]  Sheena E. Radford,et al.  Structural and mechanistic basis of immunity toward endonuclease colicins , 1999, Nature Structural Biology.

[5]  Wei Li,et al.  Crystal structure of a stable dimer reveals the molecular basis of serpin polymerization , 2008, Nature.

[6]  D. A. Low,et al.  Contact-Dependent Growth Inhibition Causes Reversible Metabolic Downregulation in Escherichia coli , 2009, Journal of bacteriology.

[7]  E. Willery,et al.  Beta‐helix model for the filamentous haemagglutinin adhesin of Bordetella pertussis and related bacterial secretory proteins , 2001, Molecular microbiology.

[8]  David A. Low,et al.  A widespread family of polymorphic contact-dependent toxin delivery systems in bacteria , 2010, Nature.

[9]  D. Eisenberg,et al.  Thiol-disulfide exchange in an immunoglobulin-like fold: structure of the N-terminal domain of DsbD. , 2002, Biochemistry.

[10]  Robert T Sauer,et al.  Engineering controllable protein degradation. , 2006, Molecular cell.

[11]  S. Aoki,et al.  Contact-Dependent Inhibition of Growth in Escherichia coli , 2005, Science.

[12]  S. Aoki,et al.  Contact‐dependent growth inhibition requires the essential outer membrane protein BamA (YaeT) as the receptor and the inner membrane transport protein AcrB , 2008, Molecular microbiology.

[13]  Angela M Gronenborn,et al.  Protein acrobatics in pairs--dimerization via domain swapping. , 2009, Current opinion in structural biology.

[14]  Matthias Bochtler,et al.  Crystal structure of the ββα-Me type II restriction endonuclease Hpy99I with target DNA , 2009, Nucleic acids research.

[15]  L. J. Perry,et al.  Protein production in Escherichia coli for structural studies by X-ray crystallography. , 2003, Journal of structural biology.

[16]  W. Lim,et al.  Unexpected modes of PDZ domain scaffolding revealed by structure of nNOS-syntrophin complex. , 1999, Science.

[17]  G. Kachalova,et al.  Structural analysis of the heterodimeric type IIS restriction endonuclease R.BspD6I acting as a complex between a monomeric site-specific nickase and a catalytic subunit. , 2008, Journal of molecular biology.

[18]  F. Niesen,et al.  The use of differential scanning fluorimetry to detect ligand interactions that promote protein stability , 2007, Nature Protocols.

[19]  T. Ko,et al.  The crystal structure of the DNase domain of colicin E7 in complex with its inhibitor Im7 protein. , 1999, Structure.

[20]  M. Inouye,et al.  Toxin-antitoxin systems in bacteria and archaea. , 2011, Annual review of genetics.

[21]  Molecular basis of inhibition of the ribonuclease activity in colicin E5 by its cognate immunity protein. , 2006, Journal of molecular biology.

[22]  Colin Kleanthous,et al.  Colicin Biology , 2007, Microbiology and Molecular Biology Reviews.

[23]  Marc Graille,et al.  Structural inhibition of the colicin D tRNase by the tRNA‐mimicking immunity protein , 2004, The EMBO journal.

[24]  G. Moore,et al.  Competitive recruitment of the periplasmic translocation portal TolB by a natively disordered domain of colicin E9 , 2006, Proceedings of the National Academy of Sciences.

[25]  Colin Kleanthous,et al.  Structure-based Analysis of the Metal-dependent Mechanism of H-N-H Endonucleases* , 2004, Journal of Biological Chemistry.

[26]  D. Eisenberg,et al.  Crystal structure of a major secreted protein of Mycobacterium tuberculosis—MPT63 at 1.5‐Å resolution , 2002, Protein science : a publication of the Protein Society.

[27]  L. Aravind,et al.  A novel immunity system for bacterial nucleic acid degrading toxins and its recruitment in various eukaryotic and DNA viral systems , 2011, Nucleic acids research.

[28]  Liisa Holm,et al.  Searching protein structure databases with DaliLite v.3 , 2008, Bioinform..

[29]  S. Aoki,et al.  The toxin/immunity network of Burkholderia pseudomallei contact‐dependent growth inhibition (CDI) systems , 2012, Molecular microbiology.

[30]  S. Aoki,et al.  Identification of Functional Toxin/Immunity Genes Linked to Contact-Dependent Growth Inhibition (CDI) and Rearrangement Hotspot (Rhs) Systems , 2011, PLoS genetics.