Structural Basis for Variations in Polo-like Kinase 1 Conformation and Intracellular Stability Induced by...

Area of Science:

• Molecular Biology
• Cell Biology
• Biochemistry

Background:

• Polo-like kinase 1 (PLK1) is crucial for cell division, featuring a kinase domain (KD) and a polobox domain (PBD) for substrate binding and localization.
• PLK1 regulation involves an autoinhibitory conformation where KD and PBD interact.
• Abbapolins, previously identified PBD-binding molecules, inhibit PLK1 substrate phosphorylation and reduce intracellular PLK1 levels.

Purpose of the Study:

• To compare the effects of abbapolins (PBD binders) and kinase domain (KD) inhibitors on PLK1 conformation and cellular activity.
• To gain insights into the distinct conformational features of PLK1 induced by PBD versus KD engagement.

Main Methods:

• Cellular thermal shift assay (CETSA) to measure ligand-induced thermal stabilization of PLK1.
• Binding measurements using full-length PLK1 and a KD inhibitor.
• Analysis of predicted structures using AlphaFold for PLK1's catalytic domain and full-length form.

Main Results:

• Abbapolins induced thermal stabilization of PLK1, indicating PBD engagement stabilizes the protein.
• KD inhibitors decreased soluble PLK1 and demonstrated a conformational change, suggesting catalytic-site binding leads to a less stable conformation.
• KD binding caused PLK1 accumulation, while PBD binding led to a loss of nuclear PLK1, highlighting contrasting cellular consequences.

Conclusions:

• KD and PBD binding induce distinct conformational changes in PLK1, impacting its stability and localization differently.
• KD inhibitors may relieve autoinhibition, potentially promoting noncatalytic functions and explaining limited clinical efficacy.
• Understanding these conformational effects is crucial for developing effective PLK1-targeting therapeutics.