Monomeric recombinant MD-2 binds toll-like receptor 4 tightly and confers lipopolysaccharide responsiveness.
In order to mediate cellular response to lipopolysaccharide (LPS), Toll-like receptor (TLR) 4 must interact with MD-2, a secreted protein. In this study, a biochemical assay was developed to demonstrate that recombinant MD-2 can interact with the extracellular portion of TLR4 in solution. The ability of MD-2 to multimerize was confirmed, and MD-1 was also shown to possess this ability. Through site-directed mutagenesis, more than two intermolecular disulfide bonds were found to stabilize the MD-2 multimer. MD-2's abilities to confer LPS responsiveness and to bind TLR4 were strongly associated functions. Remarkably, although the majority of recombinant MD-2 exists in multimeric form, monomeric MD-2 was found to preferentially bind TLR4 and to confer LPS responsiveness more efficiently than MD-2 multimers.
Re
Fabio
F
Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA. Fabio_re@dfci.harvard.edu
Strominger
Jack L
JL
D000954:Antigens, Surface / Q000737:chemistry / Q000378:metabolism*; D029721:Drosophila Proteins; D006367:HeLa Cells; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D051218:Lymphocyte Antigen 96; D008562:Membrane Glycoproteins / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012996:Solutions; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors
D000954:Antigens, Surface; D029721:Drosophila Proteins; C495377:LY96 protein, human; D008070:Lipopolysaccharides; D051218:Lymphocyte Antigen 96; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D012996:Solutions; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors