Synthesis and biochemical characterization of a photoactivatable, iodinatable, cleavable bacterial lipopolysaccharide derivative.
A method for the synthesis of a photoactivatable, iodinatable, and thiol-cleavable derivative of bacterial lipopolysaccharide (LPS) is described using sulfosuccinimidyl 2-(p-azidosalicylamido)-1,3'-dithiopropionate. The method described is applicable to LPS from both smooth and rough bacteria. Evidence is presented that the coupling reaction occurs primarily to phosphoethanolamine residues localized to the inner core region of the LPS. Radioiodination of the derivatized LPS results in a product with a specific activity of 1.8-2.5 microCi/micrograms. Experiments comparing the activity of native and derivatized S-form LPS suggest that the synthesis has not introduced major alterations in the biological properties of the LPS. The feasibility of this derivatized LPS as a molecular probe to investigate LPS binding targets in biological systems is suggested by experiments showing ultraviolet light-dependent cross-linking, thiol-dependent cleavage, and subsequent transfer of radioiodine to both monoclonal anti-LPS antibody and bovine serum albumin. The latter interaction has been demonstrated to be highly selective in protein mixtures containing serum albumin in solution with LPS.
D000656:Amoeba / Q000187:drug effects; D000818:Animals; D000911:Antibodies, Monoclonal; D001386:Azides / Q000138:chemical synthesis* / Q000378:metabolism; D001681:Biological Assay; D055598:Chemical Phenomena; D002621:Chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D008070:Lipopolysaccharides / Q000138:chemical synthesis* / Q000276:immunology; D010782:Photolysis; D013388:Succinimides / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism; D014466:Ultraviolet Rays
C047258:2-(4-azidosalicylamido)-1,3'-dithiopropionate-lipopolysaccharide; D000911:Antibodies, Monoclonal; D001386:Azides; D008070:Lipopolysaccharides; D013388:Succinimides; C047257:sulfosuccinimidyl 2-(4-azidosalicylamido)ethyl-1,3'-dithiopropionate; D013936:Thymidine