Molecular Dynamics Simulation Reveals the Structural Basis Underlying Reverse Transcriptase Activity by Human DNA polymerase 𝜼

Jeanpierre Fuente¹, Mustapha Olatunji¹, Pawlos Tsegay¹, Prem Chapagain^3,4 and Yuan Liu^1,2,4

1Biochemistry Ph.D. Program, Florida International University
2Department of Chemistry and Biochemistry, Florida International University
3Department of Physics, Florida International University
4Biomolecular Sciences Institute, Florida International University

Human DNA polymerases play an essential role in maintaining genome stability and integrity by mediating DNA replication and repair. The central dogma of genetics in mammals is that DNA is synthesized by DNA polymerases using a DNA template. However, more and more studies have pointed out that repair DNA polymerases can also employ RNA as a template to synthesize DNA suggesting that RNA also plays an active role in modulating genome stability and integrity. Yet, the effects of RNA-templated DNA synthesis on genome integrity and their underlying mechanisms remain unknown. We have recently found that human DNA polymerase 𝜼 (pol 𝜼) can perform RNA-templated DNA synthesis with similar efficiency as typical DNA-templated DNA synthesis. To further understand the structural basis of the RNA-templated DNA synthesis by pol 𝜼, we simulated DNA and RNA synthesis by pol 𝜼 substrate with DNA-DNA, RNA-DNA, and DNA-DNA in the presence of Mg2+ using Visual Molecular Dynamics (VMD) and Nanoscale Molecular Dynamics (NAMD). We have identified a major distortion in the nucleic acid substrates containing an RNA template and in the pol 𝜼-substrate ternary complex. The results showed that on both RNA and DNA template, the incoming deoxyribonucleotide adopted a similar conformation leading to efficient catalysis of RNA-templated DNA synthesis. Moreover, shifts in the palm, thumb, and little finger domain of pol 𝜼 were noted when templated with RNA versus when templated with DNA. The results indicate that pol 𝜼 adopted notable structural changes in the thumb, palm, and little finger domains to accommodate the unique conformation of the RNA template achieving efficient DNA synthesis activities. Our studies have provided new insights into the molecular mechanisms by which the reverse transcriptase activity of DNA polymerases modulate genome stability and viral infection. Our results will open a new avenue for developing novel targets for the treatment of RNA-related diseases.