Crystal twinning of human MD-2 recognizing endotoxin cores of lipopolysaccharide

Area of Science:

• Biochemistry
• Structural Biology
• Crystallography

Background:

• MD-2 protein forms a complex with Toll-like receptor 4, recognizing bacterial lipopolysaccharide (LPS).
• LPS-induced immune responses can lead to septic shock.
• Crystal twinning complicates X-ray diffraction analysis for structure determination.

Purpose of the Study:

• To overcome crystal twinning challenges in saccharide-trimmed human MD-2.
• To determine the crystal structure of MD-2.
• To elucidate the structure of the MD-2 complex with antiendotoxic lipid IVa.

Main Methods:

• Crystallization of saccharide-trimmed human MD-2 in a tetragonal form.
• Optimization of cryoprotectant to transform twinned domains into a single domain.
• Structure determination using the multiple isomorphous replacement method.

Main Results:

• Crystal twinning was identified in the tetragonal form of MD-2.
• A cryoprotectant optimization strategy successfully yielded single-domain crystals.
• The crystal structure of MD-2 in space group P4(1)2(1)2 was determined.
• The structure of the MD-2-lipid IVa complex was also successfully elucidated.

Conclusions:

• Overcoming crystal twinning is crucial for determining the structure of MD-2.
• The determined structures provide insights into MD-2's role in LPS recognition and immune response.
• This work facilitates further research into potential anti-endotoxic therapies.