Oral Presentation 2014 International Biophysics Congress

Crystal structure determination of the integral membrane diacylglycerol kinase (#29)

Dianfan Li 1 , Joseph A. Lyons 1 , Valerie E. Pye 2 , Lutz Vogeley 1 , David Aragao 3 , Colin P. Kenyon 4 , Syed T.A. Sha 1 , Christine Doherty 1 5 , Margaret Aherne 1 , Martin Caffrey 1
  1. School of Biochemistry and Immunology & School of Medicine, Trinity College Dublin, Dublin, Ireland
  2. Clare Hall Laboratories, Cancer Research UK, Hertfordshire, UK
  3. Australian Synchrotron, Clayton, VIC, Australia
  4. Biosciences, Council for Scientific and Industrial Research, Pretoria, South Africa
  5. Kennedy Institute of Rheumatology, University of Oxford, London, UK

Diacylglycerol kinase (DgkA) catalyses the ATP-dependent phosphorylation of diacylglycerol to phosphatidic acid for use in shuttling water-soluble components to membrane-derived oligosaccharide and lipopolysaccharide in the cell envelope of Gram-negative bacteria. For half a century, this 121 residue kinase has served as a model for investigating membrane protein enzymology, folding, assembly and stability. Here we present a case study where the use of non-standard lipids for the in meso crystallization methodology 1-4 successfully yielded the tridimensional structure of DgkA5 . Considerations are made on data collections strategies and difficulties during structure determination. The crystal structures rationalize extensive biochemical and biophysical data on the enzyme. They are,however, at variance with a published solution NMR model in that domain swapping, a key feature of the solution form, is not observed in the crystal structures.

  1. Höfer N, Aragão D, et al. (2011), “Membrane Protein Crystallization in Lipidic Mesophases. Hosting lipid affects on the crystallization and structure of a transmembrane peptide”, Cryst Growth Des., 11:1182-1192.
  2. Li D, Lee J, Caffrey M. (2011), “Crystallizing Membrane Proteins in Lipidic Mesophases. A Host Lipid Screen”, Cryst Growth Des,. 11:530-537.
  3. Rosenbaum DM, Zhang C, et al.., "Structure and function of an irreversible agonist-β(2) adrenoceptor complex.", Nature, 469:236-240
  4. Lyons JA, Aragão D, et al. "Structural insights into electron transfer in caa3-type cytochrome oxidase", Nature, 487:514-518
  5. Li D, Lyons JA, et al. (2013), “Crystal structure of the integral membrane diacylglycerol kinase”, Nature, 497:521-524