SNP-Associated Substitutions of Amino Acid Residues in the dNTP Selection Subdomain Decrease Polβ Polymerase Activity

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• DNA polymerase β (Polβ) is crucial for genome stability and base excision repair (BER).
• Single-nucleotide polymorphisms (SNPs) in the POLB gene can lead to altered Polβ enzymatic properties and are linked to cancer.
• Specific amino acid positions (Gly-274, Gly-290, Arg-333) are implicated in Polβ function and have been observed in tumors.

Purpose of the Study:

• To investigate the functional impact of naturally occurring polymorphic variants G274R, G290C, and R333W of DNA polymerase β.
• To elucidate how these specific amino acid substitutions affect Polβ's enzymatic activity, DNA binding, and polymerization efficiency.

Main Methods:

• Kinetic analyses were performed to quantify the enzymatic activity of the Polβ variants.
• Molecular dynamics simulations were employed to examine the structural and dynamic changes induced by the amino acid substitutions.
• Assays for gap-filling, primer elongation, and deoxynucleotide triphosphate binding were conducted.

Main Results:

• All three polymorphic variants (G274R, G290C, R333W) exhibited reduced polymerase activity.
• The G274R and R333W substitutions significantly impaired gap-filling and primer elongation capabilities.
• These variants showed decreased deoxynucleotide triphosphate binding affinity and lower polymerization constants, linked to altered local protein structures.

Conclusions:

• The studied Polβ variants (G274R, G290C, R333W) possess diminished enzymatic functions.
• These functional deficits suggest a potential role in the accumulation of unrepaired DNA damage.
• The findings highlight the importance of specific Polβ residues in maintaining genome integrity and their implications in disease.