Premium
Lipopolysaccharide‐protein interactions: Determination of dissociation constants by affinity electrophoresis
Author(s) -
Borneleit Petra,
Blechschmidt Bernd,
Kleber HansPeter
Publication year - 1989
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150101209
Subject(s) - chemistry , dissociation constant , dissociation (chemistry) , chromatography , acinetobacter calcoaceticus , chloroform , affinity electrophoresis , polyacrylamide gel electrophoresis , ligand (biochemistry) , affinity chromatography , biochemistry , organic chemistry , enzyme , receptor , acinetobacter , antibiotics
An affinity electrophoresis system is described to allow determination of dissociation constants of lipopolysaccharide (LPS)‐protein complexes. The LPS ligand is incorporated into polyacrylamide gels by addition to the polyacrylamide‐ N, N ′‐methyl‐enebisacrylamide polymerization mixture. Quantitative evaluation revealed formation of immobile protein‐ligand complexes. The method was applied both to R‐ and S‐ form LPS from Acinetobacter calcoaceticus . For a heat‐modifiable outer membrane protein with M r 18 000 from strain 69V the dissociation constant was determined to be 0.5 mM (EDTA‐salt extracted R‐LPS) and 0.3 mM (phenol‐chloroform‐petrolether extracted R‐LPS). In comparison, for another A. calcoaceticus strain, CCM 5593, a higher dissociation constant of 1.0 mM (phenol‐chloroform‐petrolether extracted R‐LPS) ‐ indicative of lower affinity ‐ was obtained. When S‐LPS from A. calcoaceticus 69V was incorporated into the affinity gels, a dissociation constant of 0.02 m M was determined which indicates much stronger interactions than those exerted by R‐LPS forms.