Abstract
Most Gram-negative bacteria are surrounded by a glycolipid called lipopolysaccharide (LPS), which forms a barrier to hydrophobic toxins and, in pathogenic bacteria, is a virulence factor. During LPS biosynthesis, a membrane-associated glycosyltransferase (LpxB) forms a tetra-acylated disaccharide that is further acylated to form the membrane anchor moiety of LPS. Here we solve the structure of a soluble and catalytically competent LpxB by X-ray crystallography. The structure reveals that LpxB has a glycosyltransferase-B family fold but with a highly intertwined, C-terminally swapped dimer comprising four domains. We identify key catalytic residues with a product, UDP, bound in the active site, as well as clusters of hydrophobic residues that likely mediate productive membrane association or capture of lipidic substrates. These studies provide the basis for rational design of antibiotics targeting a crucial step in LPS biosynthesis.
| Original language | English (US) |
|---|---|
| Article number | 377 |
| Journal | Nature communications |
| Volume | 9 |
| Issue number | 1 |
| DOIs | |
| State | Published - Dec 1 2018 |
Bibliographical note
Funding Information:We thank Surajit Banerjee for his outstanding assistance during diffraction data collection at the Advanced Photon Source. This work is based upon research conducted at the Northeastern Collaborative Access Team beamlines, which are funded by the National Institute of General Medical Sciences from the National Institutes of Health (P41 GM103403). The Pilatus 6M detector on 24-ID-C beam line is funded by a NIH-ORIP HEI grant (S10 RR029205). This research used resources of the Advanced Photon Source, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract No. DE-AC02-06CH11357. We thank Louis E. Metzger IV for his consultation on measuring the activity of LpxB including the purification of UDP-DAG and lipid X, and the proper method of charring TLC plates for visualization of lipids. We thank Jayakanth Kankanala and Zhengquiang Wang for their help with mass spectrometry. UCSF Chimera was utilized to generate 3D structure figures and for protein structure comparisons. Chimera is developed by the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco (supported by NIGMS P41-GM103311). This work was supported by NIH grant GM118047 to H.A.
Funding Information:
We thank Surajit Banerjee for his outstanding assistance during diffraction data collection at the Advanced Photon Source. This work is based upon research conducted at the Northeastern Collaborative Access Team beamlines, which are funded by the National Institute of General Medical Sciences from the National Institutes of Health (P41 GM103403). The Pilatus 6M detector on 24-ID-C beam line is funded by a NIH-ORIP HEI grant (S10 RR029205). This research used resources of the Advanced Photon Source, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract No. DE-AC02- 06CH11357. We thank Louis E. Metzger IV for his consultation on measuring the activity of LpxB including the purification of UDP-DAG and lipid X, and the proper method of charring TLC plates for visualization of lipids. We thank Jayakanth Kankanala and Zhengquiang Wang for their help with mass spectrometry. UCSF Chimera was utilized to generate 3D structure figures and for protein structure comparisons. Chimera is developed by the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco (supported by NIGMS P41-GM103311). This work was supported by NIH grant GM118047 to H.A.
Publisher Copyright:
© 2018 The Author(s).