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Apolipophorin III ‐ lipopolysaccharide interaction using tyrosine fluorescence
Author(s) -
Pratt Cindy C,
Leon Leonardo J,
Vasquez Leslie J,
Weers Paul M M
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a913-d
Subject(s) - lipopolysaccharide , lipid a , tyrosine , biochemistry , binding site , residue (chemistry) , glucosamine , fluorescence , chemistry , biophysics , biology , endocrinology , physics , quantum mechanics
Apolipophorin III (apoLp‐III) is a model exchangeable apolipoprotein, and plays a crucial role in lipid transport processes. ApoLp‐III also interacts with lipopolysaccharides (LPS), and may function as a pattern recognition protein. While the lipid A portion of LPS is involved in the binding interaction, apoLp‐III shows a strong interaction with the LPS carbohydrate portion. To study the LPS binding site in more detail, binding studies were performed using the intrinsic Tyr fluorescence properties of apoLp‐III of Galleria mellonella. The unique Tyr‐142 residue is highly quenched in the unbound state, but becomes unquenched when bound to LPS. We showed that this unquenched state exists at pH 7‐9, while at a pH < 7 Tyr becomes unquenched. Therefore all binding assays were performed in phosphate buffered saline at pH 7.4. The binding of apoLp‐III to a truncated version of LPS (Rd), which lacks the O‐antigen, was increased 20‐fold compared to LPS, indicating that core sugars play a key‐role in binding. Hexoses, found in the core of LPS, were incubated with LPS to compete for apoLp‐III binding. A 60% decrease in Tyr fluorescence was observed with glucose and N‐acetyl‐glucosamine. This indicates that the core sugars of LPS are primarily responsible for apoLp‐III binding interaction. This work was supported by grants from the National Institutes of Health (R15 HL077135 and S06 GM 063119‐05).