Expression, purification and crystallization of a membrane protein from <em>Neisseria meningitides</em> - LptA — ASN Events
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  3. Expression, purification and crystallization of a membrane protein from Neisseria meningitides - LptA

Expression, purification and crystallization of a membrane protein from Neisseria meningitides - LptA (#214)

Anandhi Anandan 1 , Susannah Piek 1 , Isabel Moraes 2 , Charlene Kahler 1 , Alice Vrielink 1
  1. Univesity of Western Australia, Crawley, WA, Australia
  2. Membrane Protein Laboratory, Didcot, United Kingdom

Neisseria meningitidis (NM), an obligate human pathogen, is the causative agent of meningitis and septic shock. The Neisserial endotoxin is a major virulence factor that induces high levels of pro-inflammatory mediators 4 .

The endotoxin is composed of a Lipid A moiety, which can be modified through the addition of phosphoethanolamine (PEA) by the enzyme Lipooligosaccharide phophoethanolamine transferase A (LptA) 1 . This modification plays an important role in increasing the ability of the bacterium to attach to host cells 2  and their resistance to the host immune system 3 . It has been proposed that NMLptA catalysis the addition of PEA from phosphatidylethanolamine to lipid A. Understanding the catalysis and substrate binding of this enzyme may enable future development of small molecule inhibitors for controlling Neisserial infections. LptA has been predicted to be an integral membrane protein consisting of a transmembrane domain and a periplasmic soluble globular domain. The structure of the soluble globular domain was solved earlier in our laboratory. The truncated structure of LptA, its substrate and ligand binding experiments of the soluble domain indicate that the transmembrane domain may play a major role in catalysis of the enzyme.  Hence structural studies of full-length NMLptA have been undertaken. 

 NMLptA (full length) has been successfully expressed in a recombinant E. coli expression system, purified to homogeneity in the presence of a detergent and crystallized. A preliminary X-ray diffraction data set to 3Å resolution has been collected from full-length NMLptA crystals.           

  1. Cox A. D., Wright, J. C., Li J., Hood, D. W., Moxon, E. R., Richards, J. C. (2003). Journal of Bacteriology 185, 3270 - 3277.
  2. Takahashi, H., Carlson, R. W., Muszynski, A., Choudhury, B., Kim, K. S., Stephens, D. S., Watanabe, H. (2008). Infection and Immunity 76, 5777 - 5789.
  3. Tzeng, Y. L., Ambrose, K. D., Zughaier, S., Zhou, X., Miller, Y. K., Shafer, W. M., Stephens, D. S. (2005). Journal of Bacteriology 187, 5387 - 5396.
  4. Yazdankhah, S. P., Caugant, D. A. (2004). Journal of Medical Microbiology 53, 821 - 832.