Molecular Effects of Site-Specific Phosphate-Methylated Primer on the Structure and Motions of Taq DNA Polymerase

Computational and structural biotechnology journal(2023)

引用 0|浏览7
暂无评分
摘要
Polymerase chain reaction (PCR) is a powerful molecular biology assay for gene detection and quantification. Conventional DNA primers for PCR often suffer from poor sensitivity in specific gene detection. Recently, oligonucleotides containing methyl phosphotriester (MPTE-DNA) have been developed with enhanced DNA hybridization and improved gene detection sensitivity. Yet, site-specific MPTE-modifications on DNA primers have been reported to affect PCR amplification efficiencies while the detailed mechanism remains elusive. Here, we utilized molecular dynamics (MD) simulation to examine the effects of site-specific MPTE-modified primers on the structure and motions of DNA/Taq polymerase complexes. All tested MPTE-DNA/Taq complexes exhibited RMSD values below 2 Å, indicating insignificant effects of all methylation sites on the complex stability. The energetic and hydrogen-bonding analyses suggest minor effects of methylation at t-3, t-4, t-6, and t-7 positions on the DNA−Taq interaction. Principal component analyses further reveal that only t-3, and t-7 methylations preserve the motions of the Taq thumb domain. The site-specific methylation affects the interaction between DNA and the surrounding protein residues, resulting in allosteric-like effects on the DNA/Taq complex. The MD data complement the best experimentally observed PCR efficacies for the t-3 and t-7 positions among all tested MPTE-primers. The unveiled molecular insights can benefit the design of novel PCR primers for improving genetic testing platforms.
更多
查看译文
关键词
PCR,Primer,Methyl phosphotriester,DNA polymerase,Molecular dynamics
AI 理解论文
溯源树
样例
生成溯源树,研究论文发展脉络
Chat Paper
正在生成论文摘要