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Lipopolysaccharide binding to the periplasmic protein LptA. Protein Sci 2017 Aug;26(8):1517-1523

Date

04/19/2017

Pubmed ID

28419595

Pubmed Central ID

PMC5521551

DOI

10.1002/pro.3177

Scopus ID

2-s2.0-85018394516 (requires institutional sign-in at Scopus site)   24 Citations

Abstract

Lipopolysaccharide (LPS) and the periplasmic protein, LptA, are two essential components of Gram-negative bacteria. LPS, also known as endotoxin, is found asymmetrically distributed in the outer leaflet of the outer membrane of Gram-negative bacteria such as Escherichia coli and plays a role in the organism's natural defense in adverse environmental conditions. LptA is a member of the lipopolysaccharide transport protein (Lpt) family, which also includes LptC, LptDE, and LptBFG2 , that functions to transport LPS through the periplasm to the outer leaflet of the outer membrane after MsbA flips LPS across the inner membrane. It is hypothesized that LPS binds to LptA to cross the periplasm and that the acyl chains of LPS bind to the central pocket of LptA. The studies described here are the first to comprehensively characterize and quantitate the binding of LPS by LptA. Using site-directed spin-labeling electron paramagnetic resonance (EPR) spectroscopy, data were collected for 15 spin-labeled residues in and around the proposed LPS binding pocket on LptA to observe the mobility changes caused by the presence of exogenous LPS and identify the binding location of LPS to LptA. The EPR data obtained suggest a 1:1 ratio for the LPS:LptA complex and allow the first calculation of dissociation constants for the LptA-LPS interaction. The results indicate that the entire protein is affected by LPS binding, the N-terminus unfolds in the presence of LPS, and a mutant LptA protein unable to form oligomers has an altered affinity for LPS.

Author List

Schultz KM, Lundquist TJ, Klug CS

Authors

Candice S. Klug PhD Professor in the Biophysics department at Medical College of Wisconsin
Kathryn M. Schultz Research Scientist I in the Biophysics department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Binding Sites
Carrier Proteins
Cloning, Molecular
Electron Spin Resonance Spectroscopy
Escherichia coli
Escherichia coli Proteins
Gene Expression
Kinetics
Lipopolysaccharides
Models, Molecular
Mutation
Periplasm
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Protein Multimerization
Protein Structure, Tertiary
Recombinant Proteins
Spin Labels