Human Polβ Natural Polymorphic Variants G118V and R149I Affects Substate Binding and Catalysis

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• DNA polymerase beta (Polβ) is crucial for DNA repair, particularly in the base excision repair pathway.
• Polβ's function is vital for maintaining genomic stability; defects are linked to diseases like cancer and premature aging.
• Single-nucleotide polymorphisms in the POLB gene can alter Polβ function, but their specific effects are often unclear.

Purpose of the Study:

• To investigate the functional consequences of two specific human Polβ polymorphic variants (G118V and R149I) on DNA repair.
• To determine how these variants affect Polβ's interaction with gapped DNA and its catalytic efficiency.

Main Methods:

• Site-directed mutagenesis was used to create the G118V and R149I Polβ variants.
• DNA-binding assays were performed to assess the affinity of wild-type and variant Polβ for gapped DNA and dATP.
• Enzyme kinetics were measured to determine the catalytic rates of the variants compared to the wild-type enzyme.

Main Results:

• Both G118V and R149I variants exhibited reduced binding affinity for gapped DNA compared to wild-type Polβ.
• The polymorphic variants also showed a weakened binding affinity for the substrate dATP.
• The G118V variant significantly impaired Polβ's ability to fill DNA gaps and markedly reduced its catalytic rate.

Conclusions:

• The studied Polβ polymorphic variants (G118V and R149I) negatively impact DNA repair by reducing DNA-binding affinity and catalytic efficiency.
• These findings suggest that certain POLB gene polymorphisms can compromise base excision repair, potentially contributing to increased mutation frequencies and associated diseases.