Identification of single amino acid residues essential for the binding of lipopolysaccharide (LPS) to LPS binding protein (LBP) residues 86-99 by using an Ala-scanning library.
Lipopolysaccharide binding protein (LBP) is a 60 kDa acute phase glycoprotein capable of binding to LPS of Gram-negative bacteria and facilitating its interaction with cellular receptors. This process is thought to be of great importance in systemic inflammatory reactions such as septic shock. A peptide corresponding to residues 86-99 of human LBP (LBP86-99) has been reported to bind specifically with high affinity the lipid A moiety of LPS and to inhibit the interaction of LPS with LBP. We identified essential amino acids in LBP86-99 for binding to LPS by using a peptide library corresponding to the Ala-scanning of human LBP residues 86-99. Amino acids Trp91 and Lys92 were indispensable for peptide-LPS interaction and inhibition of LBP-LPS binding. In addition, several alanine-substituted synthetic LBP-derived peptides inhibited LPS-LBP interaction. Substitution of amino acids Arg94, Lys95 and Phe98 by Ala increased the inhibitory effect. The mutant Lys95 was the most active in blocking LPS binding to LBP. These findings emphasize the importance of single amino acids in the LPS binding capacity of small peptides and may contribute to the development of new drugs for use in the treatment of Gram-negative bacterial sepsis.
Reyes
O
O
Division of Physical Chemistry, Center for Genetic Engineering and Biotechnology, Havana, Cuba. oreyes@cigb.edu.cu
Vallespi
M G
MG
Garay
H E
HE
Cruz
L J
LJ
González
L J
LJ
Chinea
G
G
Buurman
W
W
Araña
M J
MJ
D000209:Acute-Phase Proteins; D000409:Alanine / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics*; D001665:Binding Sites; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008562:Membrane Glycoproteins; D016296:Mutagenesis; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D000099231:Lipopolysaccharide-Binding Protein
D000209:Acute-Phase Proteins; D000409:Alanine; D002352:Carrier Proteins; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D019151:Peptide Library; D010455:Peptides; D000099231:Lipopolysaccharide-Binding Protein