G-quadruplex-proximity protein labeling based on peroxidase activity.

Area of Science:

• Biochemistry
• Molecular Biology
• Chemical Biology

Background:

• Peroxidase-based protein labeling is crucial for studying protein interactions.
• G-quadruplex (G4) structures can modulate enzyme activity.
• TERRA, a G4 structure, has potential roles in gene regulation.

Purpose of the Study:

• To develop a novel proximity labeling method utilizing a hemin-parallel G-quadruplex (G4) complex.
• To investigate the enhanced peroxidase activity of hemin when bound to a parallel G4 structure.
• To demonstrate the utility of this method for labeling RNA-binding proteins in a cellular context.

Main Methods:

• Formation of a hemin-parallel G-quadruplex (G4) complex.
• Utilizing an N-methyl luminol derivative for tyrosine labeling.
• Assessing labeling efficiency in proximity to the hemin-G4 complex.
• Application of the method to HeLa cell lysates to label RNA-binding proteins.

Main Results:

• The hemin-parallel G4 complex significantly enhanced peroxidase activity.
• Proximity to the hemin-G4 complex accelerated the tyrosine labeling reaction.
• The TERRA-hemin complex successfully activated labeling of multiple RNA-binding proteins, including heterogeneous nuclear ribonucleoproteins.
• Labeling was demonstrated in a complex biological matrix (HeLa cell lysate).

Conclusions:

• Hemin-parallel G4 complexes offer a powerful platform for enhanced peroxidase-mediated proximity labeling.
• This method provides a sensitive approach for identifying and characterizing RNA-binding proteins.
• The TERRA-hemin complex represents a promising tool for biochemical and cell biology research.