Phosphorylation promotes the endonuclease-like activity of human centrin 2

Area of Science:

• Biochemistry
• Molecular Biology
• Protein Chemistry

Background:

• Centrin, an EF-hand calcium-binding protein, plays a role in DNA repair.
• Post-translational modification, specifically phosphorylation, is a key regulatory mechanism for centrin in vivo.
• Understanding centrin's function in nucleotide excision repair (NER) is crucial for cellular processes.

Purpose of the Study:

• To investigate the impact of phosphorylation on the endonuclease-like activity of human centrin 2 (HsCen2).
• To explore the influence of terbium ions (Tb3+) on HsCen2 activity and DNA binding.
• To elucidate the regulatory role of phosphorylation in centrin's interaction with DNA.

Main Methods:

• Spectroscopy techniques were employed to analyze protein-ligand interactions.
• Gel electrophoresis was used to assess DNA binding.
• Molecular docking simulations provided insights into binding mechanisms.
• Experiments were conducted in Hepes buffer at pH 7.4.

Main Results:

• Phosphorylation decreased Tb3+ binding to HsCen2.
• Phosphorylation enhanced DNA binding affinity to HsCen2.
• HsCen2 exhibited improved endonuclease-like activity upon phosphorylation.
• Tb3+ ions promoted both DNA binding and endonuclease-like activity of HsCen2, irrespective of phosphorylation status.

Conclusions:

• Phosphorylation significantly modulates HsCen2's interaction with DNA and Tb3+.
• The findings highlight phosphorylation as a critical regulator of HsCen2's protein-driven functions in DNA repair.
• Tb3+ acts as a beneficial cofactor for HsCen2's DNA binding and enzymatic activity.