NMR of Membrane Proteins in Membranes

Francesca M. MarassiBurnham Institute for Medical Research

NIGMS Roadmap for Membrane Protein StructuresMarch 27 2009, UC San Francisco

• E. coli OmpX.

• Y. Pestis Ail.

• M. tuberculosis OmpATb.

Complementary approaches to membrane proteins: β-barrel bacterial outer membrane proteins.

solution NMRin micelles

solid-state NMRin lipid bilayers

X-ray diffractionIn crystals

Marassi and Opella (2000) J. Magn. Reson. 144:150Wang et al. (2000) J. Magn. Reson. 144:162Marassi (2001) Biophys. J. 80:994

Ail/Lom bacterial outer membrane proteins.

• adhesion to host epithelial cells• resistance to complement-mediated killing • pathogenesis

• NMR in membranes• Barrel tilt• Structure determination

Vogt & Schulz (1999) Structure 7:1301Fernandez et al. (2004) J. Mol. Bio. 336:1211

Ail/Lom bacterial outer membrane proteins: E. coli OmpX.

Mahalakshmi et al. (2007) BBA Biomembranes, 1768:3216

SDS SDS14-O-PC 14-O-PC

E. coli OmpX in magnetically oriented phospholipid bicelles.

Mahalakshmi et al. (2007) BBA Biomembranes, 1768:3216

H2O

D2O

E. coli OmpX in magnetically oriented phospholipid bicelles.

Mahalakshmi and Marassi (2008) Biochemistry, 47:6531

assignrefine

back-calculate

Yersinia Pestis Ail outer membrane protein: production.

Yersinia Pestis Ail outer membrane protein: refolding.

Yersinia Pestis Ail outer membrane protein in micelles.

Mycobacterium tuberculosis outer membrane protein OmpATb.

Mycobacterium tuberculosis outer membrane protein OmpATb.

Search of FFAS Server yields homology toBON (Bacterial OsmY and Nodulation) domain• 60 residues• αβ predicted fold• osmotic shock protection• interaction with membrane lipids

Yeats and Bateman (2003) TRENDS Biochem. Sci., 28:352

Thank YouOmpXR. Mahalakshmi (Burnham)Cecile Loudet (Burnham)Georg Schulz (A.L.U., Freiburg)

AilCandace Rypien (Burnham)Leigh Plesniak (U. San Diego)Greg Plano (U. Miami)

OmpATbPeter Teriete (Burnham)Adrian Kolodzik (Burmham)Michael Niederweis (U. Alabama, Birmingham)

TBBPC bicellesErick Dufourc (U. Bordeaux)

NIH/NIGMSRoadmap for Membrane Protein Structures