The structure of the ternary Eg5-ADP-ispinesib complex.

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• The human kinesin Eg5 is crucial for forming bipolar spindles during mitosis.
• Eg5 inhibition leads to mitotic arrest and apoptosis, making it a promising cancer drug target.
• Ispinesib is an advanced Eg5-targeting agent with demonstrated antitumor activity, currently in Phase II clinical trials.

Purpose of the Study:

• To determine the crystal structure of the Eg5 motor domain in complex with the inhibitor ispinesib.
• To elucidate the binding interactions between Eg5 and ispinesib at an atomic level.
• To provide structural insights that can aid in the development of novel Eg5-targeting cancer therapeutics.

Main Methods:

• X-ray crystallography was used to determine the structure of the Eg5-ispinesib complex.
• Kinetic and thermodynamic binding assays were performed to characterize the interaction.
• Advanced data processing and refinement techniques were employed to overcome crystallographic challenges.

Main Results:

• The crystal structure of the Eg5 motor domain bound to ispinesib was determined.
• Ispinesib was found to bind in an induced-fit pocket, forming extensive hydrophobic interactions with Eg5.
• The study addressed and overcame challenges related to pseudo-merohedral twinning and non-crystallographic symmetry.

Conclusions:

• The determined structure provides a detailed atomic view of Eg5 inhibition by ispinesib.
• These findings offer valuable structural information for the rational design of future Eg5 inhibitors.
• The study overcomes previous limitations in structural data for Eg5-targeting agents.