Hydrolytically active Eu(III) and Ce(IV) EF-hand peptides.

Area of Science:

• Biochemistry
• Molecular Biology
• Bioinorganic Chemistry

Background:

• The engrailed homeodomain contains a helix-turn-helix motif for DNA binding.
• EF-hand motifs are known for metal ion binding.
• Combining these motifs could create novel metallopeptides with catalytic activity.

Purpose of the Study:

• To design and characterize a chimeric peptide (P4) integrating an EF-hand metal-binding loop with the engrailed homeodomain's DNA binding motif.
• To investigate the metal-binding properties and catalytic activity of P4 towards DNA and phosphate esters.
• To explore the structural and functional consequences of metal binding on the chimeric peptide.

Main Methods:

• Peptide design and synthesis of the chimeric peptide (P4).
• Lanthanide binding studies using techniques like Circular Dichroism (CD) spectroscopy.
• Assays for DNA cleavage and hydrolysis of model phosphate esters (bisnitrophenyl phosphate).

Main Results:

• The chimeric peptide P4 successfully binds lanthanides (Eu(III) and Ce(IV)) with moderate affinities.
• P4, in the presence of bound lanthanides, efficiently cleaves supercoiled DNA and model phosphate esters.
• Metal binding enhances P4's structure, though the peptide remains flexible.
• Catalytic activity is dependent on the metallopeptide, not the peptide or metal alone.

Conclusions:

• Designed DNA-binding EF-hand peptides can deliver metal ions to DNA for catalysis.
• Efficient catalytic activity can be achieved even with flexible metallopeptide structures.
• This work presents a novel approach to creating metalloenzymes for DNA modification and catalysis.