Influence of the lipid membrane environment on structure and activity of the outer membrane protein Ail from Yersinia pestis. Biochim Biophys Acta 2015 Feb;1848(2):712-20
Date
11/30/2014Pubmed ID
25433311Pubmed Central ID
PMC4281492DOI
10.1016/j.bbamem.2014.11.021Scopus ID
2-s2.0-84915758524 (requires institutional sign-in at Scopus site) 26 CitationsAbstract
The surrounding environment has significant consequences for the structural and functional properties of membrane proteins. While native structure and function can be reconstituted in lipid bilayer membranes, the detergents used for protein solubilization are not always compatible with biological activity and, hence, not always appropriate for direct detection of ligand binding by NMR spectroscopy. Here we describe how the sample environment affects the activity of the outer membrane protein Ail (attachment invasion locus) from Yersinia pestis. Although Ail adopts the correct β-barrel fold in micelles, the high detergent concentrations required for NMR structural studies are not compatible with the ligand binding functionality of the protein. We also describe preparations of Ail embedded in phospholipid bilayer nanodiscs, optimized for NMR studies and ligand binding activity assays. Ail in nanodiscs is capable of binding its human ligand fibronectin and also yields high quality NMR spectra that reflect the proper fold. Binding activity assays, developed to be performed directly with the NMR samples, show that ligand binding involves the extracellular loops of Ail. The data show that even when detergent micelles support the protein fold, detergents can interfere with activity in subtle ways.
Author List
Ding Y, Fujimoto LM, Yao Y, Plano GV, Marassi FMAuthor
Francesca M. Marassi PhD Chair, Professor in the Biophysics department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
Amino Acid SequenceBacterial Outer Membrane Proteins
Dimyristoylphosphatidylcholine
Escherichia coli
Fibronectins
Gene Expression
Glycolipids
Humans
Inositol Phosphates
Ligands
Magnetic Resonance Spectroscopy
Membrane Lipids
Molecular Sequence Data
Nanostructures
Phosphatidylglycerols
Phospholipid Ethers
Phosphorylcholine
Protein Folding
Protein Structure, Secondary
Recombinant Proteins
Virulence Factors
Yersinia pestis
