The role of disulfide bonds in the assembly and function of MD-2

摘要: MD-2 is a secreted glycoprotein that binds to the extracellular domain of Toll-like receptor 4 (TLR4) and required for activation TLR4 by lipopolysaccharide (LPS). The protein contains seven Cys residues consists heterogeneous collection disulfide-linked oligomers. To investigate role sulfhydryls in structure function, we created 17 single multiple substitution mutants. All mutant proteins, including one totally lacking residues, were stable. SDS/PAGE analyses indicated most could participate oligomer formation no residue was oligomerization. Of substitutions, only C95S C105S failed confer LPS responsiveness on when cotransfected into cells expressing an NF-κB reporter plasmid. Surprisingly, both C95 C105 partially restored activity. Structural revealed formed intrachain disulfide bond, whereas itself produced inactive dimer. In contrast cotransfection experiments, WT conferred proteins added directly cells. Our data are consistent with model which most, possibly all lie surface stable core where they form intra- interchain bridges. These bonds produce array oligomers, some species can active complex TLR4.