Crystal structure of the TLR4-MD-2-lipopolysaccharide complex

Abstract

Together with adaptive immune system, innate immune system is indispensable part of our defense mechanism against pathogenic microbial infection. The adaptive immunity is an antigen-specific immune response, which is mediated by T and B cells, whereas innate immunity is nonspecific defense mechanisms and receptors recognize many related molecular structures called PAMPs (Pathogen-Associated Molecular Patterns). Toll-Like Receptors (TLR) family proteins play an important role in innate immune response by recognizing conserved structural patterns in diverse microbial molecules. Specially, TLR4 with its co-receptor Myeloid Differentiation factor 2 (MD-2) is responsible for LipoPolySaccharide (LPS) recognition. LPS is a major component of the outer membrane of Gram-negative bacteria and a potent inducer of innate immune response. It is extracted and transferred to TLR4-MD-2 complex by two accessory proteins, LBP (LPS-Binding Protein) and CD14. Binding of LPS to the TLR4-MD-2 complex causes a rearrangement of the extracellular part of the receptor complex, which enforce aggregation of intracellular domain signaling domain called TIR domain (TIR: Toll/Interleukin-1 Resistance). In order to understand structural basis of TLR4 activation, we undertook crystallographic studies of TLR4-MD-2 in complex with various ligand molecules. In 2007, we determined the first structure of the TLR4-MD-2 complex with bound endotoxin antagonist Eritoran. In this study, we introduced a novel technique termed “Hybrid LRR Technique” to overcome technical difficulties associated with production and crystallization of the TLR4-MD-2 complex. By using this technique, we produced a series of hybrids of human TLR4 and hagfish VLR. Among them, three hybrids named TV3, TV8 and VT3 are successfully produced and crystallized. In particular, TV3 could be crystallized together with MD-2 and Eritoran. Eritoran is a candidate drug molecule under clinical trial phase III against severe sepsis. It mimi...