Biophysical analysis of the interaction of granulysin-derived peptides with enterobacterial endotoxins.
To combat infections by Gram-negative bacteria, it is not only necessary to kill the bacteria but also to neutralize pathogenicity factors such as endotoxin (lipopolysaccharide, LPS). The development of antimicrobial peptides based on mammalian endotoxin-binding proteins is a promising tool in the fight against bacterial infections, and septic shock syndrome. Here, synthetic peptides derived from granulysin (Gra-pep) were investigated in microbiological and biophysical assays to understand their interaction with LPS. We analyzed the influence of the binding of Gra-pep on (1) the acyl chain melting of the hydrophobic moiety of LPS, lipid A, by Fourier-transform spectroscopy, (2) the aggregate structure of LPS by small-angle X-ray scattering and cryo-transmission electron microscopy, and 3) the enthalpy change by isothermal titration calorimetry. In addition, the influence of Gra-pep on the incorporation of LPS and LPS-LBP (lipopolysaccharide-binding protein) complexes into negatively charged liposomes was monitored. Our findings demonstrate a characteristic change in the aggregate structure of LPS into multilamellar stacks in the presence of Gra-pep, but little or no change of acyl chain fluidity. Neutralization of LPS by Gra-pep is not due to a scavenging effect in solution, but rather proceeds after incorporation into target membranes, suggesting a requisite membrane-bound step.
Chen
Xi
X
Division of Immunology and Transplantation Biology, Department of Pediatrics, CCSR 2105, Stanford University School of Medicine, Stanford, CA 94305, USA.
Howe
Jörg
J
Andrä
Jörg
J
Rössle
Manfred
M
Richter
Walter
W
da Silva
Ana Paula Galvão
AP
Krensky
Alan M
AM
Clayberger
Carol
C
Brandenburg
Klaus
K
D000595:Amino Acid Sequence; D000945:Antigens, Differentiation, T-Lymphocyte / Q000737:chemistry*; D001427:Bacterial Toxins / Q000737:chemistry*; D016207:Cytokines / Q000096:biosynthesis; D004731:Endotoxins / Q000737:chemistry*; D006801:Humans; D008070:Lipopolysaccharides / Q000737:chemistry*; D008081:Liposomes / Q000737:chemistry; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*
D000945:Antigens, Differentiation, T-Lymphocyte; D001427:Bacterial Toxins; D016207:Cytokines; D004731:Endotoxins; C075005:GNLY protein, human; D008070:Lipopolysaccharides; D008081:Liposomes; D010455:Peptides; C027952:salmonella toxin