As part of an international team, led by HepaRegeniX, ZoBio solved the structure of an inhibitor bound to the kinase MKK4 using a non-classical approach to structural elucidation with NMR. These efforts have recently been described in Cell article detailing the preclinical characterization and development of the clinical candidate HRX215, a first-in-class small molecule inhibitor of MKK4.  

Mitogen-Activated Protein (MAP) Kinase Kinase 4 (MKK4) is a dual specific MAP2 kinase that was recently shown to be a master regulator of liver cell (hepatocyte) regeneration. Modulation of MKK4 activity therefore has profound therapeutic implication for patients suffering from chronic or acute liver disease. Including cirrhosis of the liver, primary liver tumors and cancer metastasized to the liver.  

Efforts to obtain the structure of the inhibitor bound MKK4 kinase domain were started using X-ray crystallography. The structure obtained using this technique was of insufficient resolution to resolve the mode of binding of the inhibitor. To support structure-based inhibitor design, the strategic shift to NMR was made to obtain structural information. A close collaboration between the NMR spectroscopists and protein scientists at ZoBio enabled the use of a non-classical approach to NMR structural elucidation because conventional NMR methods yielded insufficient spectral quality.  

The method employed to finally resolve the inhibitor-bound MKK4 structure was based on selective labeling of methyl groups in the protein. These sparse spectral data were supplemented with the structural data of a close homolog and multiple rounds of point-mutant design to assign peaks and validate assignments. Together, this data package resulted in the unambiguous resolution of the structure of the MKK4-inhibitor complex.  

This study underscores the versatility of the techniques and expertise within ZoBio and demonstrates the strength of close collaboration among our scientists in solving challenging problems!