An in cellulo-derived structure of PAK4 in complex with its inhibitor Inka1

Area of Science:

• Structural Biology
• Molecular Cell Biology
• Biochemistry

Background:

• PAK4 (P21-activated kinase 4) is a key kinase in metazoan signaling pathways, downstream of Cdc42.
• Understanding PAK4's structure and function is crucial for developing targeted therapeutics.

Purpose of the Study:

• To determine the in cellulo crystal structure of human PAK4 in complex with its inhibitor, Inka1.
• To elucidate the binding mechanism of ATP and Inka1 to the PAK4 catalytic domain.

Main Methods:

• In cellulo crystallography of single mammalian cells.
• High-resolution (2.95 Å) structural determination.
• Use of Inka1-GFP for monitoring crystal formation in live cells.

Main Results:

• The crystal structure reveals detailed interactions between PAK4, ATP, and Inka1.
• The crystal lattice exhibits a hexagonal array of PAK4-PAK4 contacts with large channels.
• The crystal structure accommodates other proteins when fused to the kinase inhibitor.

Conclusions:

• The determined structure provides a detailed molecular understanding of PAK4 inhibition.
• Inka1 derivatives offer a tool for studying PAK4 inhibition in cellular and organismal models.
• This research facilitates the validation of therapeutic agents targeting PAK4.