Characterization of the apoLp-III/LPS complex: insight into the mode of binding interaction.
Apolipoproteins are able to associate with lipopolysaccharides (LPS), potentially providing protection against septic shock. To gain insight into the molecular details of this binding interaction, apolipophorin III (apoLp-III) from Galleria mellonella was used as a model. The binding of apoLp-III to LPS was optimal around 37-40 °C, close to the LPS phase transition temperature. ApoLp-III formed complexes with LPS from E. coli (serotype O55:B5) with a diameter of ~20 nm and a molecular weight of ~390 kDa, containing four molecules of apoLp-III and 24 molecules of LPS. The LPS-bound form of the protein was substantially more resistant to guanidine-induced denaturation compared to unbound protein. The denaturation profile displayed a multiphase character with a steep drop in secondary structure between 0 and 1 M guanidine-HCl and a slower decrease above 1 M guanidine-HCl. In contrast, apoLp-III bound to detoxified LPS was only slightly more resistant to guanidine-HCl induced denaturation compared to unbound protein. Analysis of size-exclusion FPLC elution profiles of mixtures of apoLp-III with LPS or detoxified LPS indicated a much weaker binding interaction with detoxified LPS compared to intact LPS. These results indicate that apoLp-III initially interacts with exposed carbohydrate regions, but that the lipid A region is required for a more stable LPS binding interaction.
Department of Chemistry and Biochemistry, California State University Long Beach, 1250 Bellflower Blvd., Long Beach, CA 90840, USA.
Merve
M
Oztug
Daisy
D
Martinon
Paul M M
PM
Weers
D000818:Animals; D001053:Apolipoproteins / Q000737:chemistry / Q000378:metabolism*; D019791:Guanidine / Q000494:pharmacology; D019476:Insect Proteins / Q000737:chemistry / Q000378:metabolism*; D007711:Klebsiella pneumoniae; D007915:Lepidoptera; D008070:Lipopolysaccharides / Q000378:metabolism*; D011485:Protein Binding; D011489:Protein Denaturation / Q000187:drug effects; D013696:Temperature
D001053:Apolipoproteins; D019476:Insect Proteins; D008070:Lipopolysaccharides; C042470:apolipophorin III; D019791:Guanidine