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PatB1 is an O-acetyltransferase that decorates secondary cell wall polysaccharides

Nature Chemical Biology volume 14, pages 79–85 (2018)Cite this article

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Abstract

O-Acetylation of the secondary cell wall polysaccharides (SCWP) of the Bacillus cereus group of pathogens, which includes Bacillus anthracis, is essential for the proper attachment of surface-layer (S-layer) proteins to their cell walls. Using a variety of pseudosubstrates and a chemically synthesized analog of SCWP, we report here the identification of PatB1 as a SCWP O-acetyltransferase in Bacillus cereus. Additionally, we report the crystal structure of PatB1, which provides detailed insights into the mechanism of this enzyme and defines a novel subfamily of the SGNH family of esterases and lipases. We propose a model for the O-acetylation of SCWP requiring the translocation of acetyl groups from a cytoplasmic source across the plasma membrane by PatA1 and PatA2 for their transfer to SCWP by PatB1.

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Figure 1: Structure of SCWP from the Bacillus cereus group and organization of genes involved in its modification.
Figure 2: Enzymatic characterization of PatB1 as a SCWP O-acetyltransferase.
Figure 3: Structure of PatB188–396.
Figure 4: Identification of catalytic and oxyanion hole residues.
Figure 5: Proposed model for the O-acetylation of SCWP in the Bacillus cereus group of pathogens.

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  • 04 December 2017

    In the version of this article initially published online, the caption of Figure 5 incorrectly stated, "Although PatA2 resembles an acetyltransferase, its function and role remain to be determined." It should read "Although PatB2 resembles an acetyltransferase, its function and role remain to be determined." The error has been corrected in the PDF and HTML versions of this article.

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Acknowledgements

We thank D. Brewer and A. Charchoglyan of the Mass Spectrometry Facility (Advanced Analysis Centre, University of Guelph) for expert technical assistance and advice, both R. Pfoh and N.C. Bamford of the Howell laboratory for their assistance in X-ray data collection, and C. Jones for technical assistance with some of the enzyme assays. P.J. Moynihan, J.M. Pfeffer, J.C. Whitney and J.T. Weadge are thanked for helpful discussions in the early stages of this research. These studies were supported in part by operating grants from the Canadian Institutes of Health Research (CIHR) to A.J.C. (TGC 114045) and P.L.H. (MOP 43998) and the Canadian Glycomics Network (http://dx.doi.org/10.13039/501100009056). J.L. was supported in part by graduate scholarships from the University of Toronto, the Ontario Graduate Scholarship Program, and CIHR. P.L.H. is the recipient of a Canada Research Chair. The National Synchrotron Light Source beam line X29A is supported by the United States Department of Energy Office of Biological and Environmental Research and the National Institutes of Health National Centre for Research Resources.

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Author notes

  1. Dustin J Little

    Present address: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, Canada

Authors and Affiliations

  1. Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada

    David Sychantha & Anthony J Clarke

  2. Program in Molecular Medicine, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada

    Dustin J Little & P Lynne Howell

  3. Department of Biochemistry, University of Toronto, Ontario, Canada

    Dustin J Little & P Lynne Howell

  4. Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia, USA

    Robert N Chapman & Geert-Jan Boons

  5. Photon Science Division, Brookhaven National Laboratory, Upton, New York, USA

    Howard Robinson

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Contributions

D.S. conceived all experiments and performed all gene cloning, site-directed mutagenesis and expression experiments; purification of recombinant proteins; enzyme kinetic and MS analyses; crystallization of proteins; collection and analysis of crystallographic data; and solving of crystal structures; and made structural figures and tables; D.J.L. and P.L.H. oversaw the collection and analysis of crystallographic data; R.N.C. and G.-J.B. synthesized the SCWP model compound, MGG, and performed all NMR experiments; H.R. collected crystallographic data; A.J.C. helped conceive the cloning, protein purification, kinetic and MS experiments, analyzed data and oversaw the work of D.S. D.S. and A.J.C. wrote the paper with contributions from all authors.

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Correspondence to Anthony J Clarke.

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Supplementary Results, Supplementary Tables 1–4 and Supplementary Figures 1–18 (PDF 9039 kb)

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Supplementary Note 1

Synthesis of MGG (PDF 602 kb)

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Sychantha, D., Little, D., Chapman, R. et al. PatB1 is an O-acetyltransferase that decorates secondary cell wall polysaccharides. Nat Chem Biol 14, 79–85 (2018). https://doi.org/10.1038/nchembio.2509

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