GC-Content Dependence of Elastic and Overstretching Properties of DNA:RNA Hybrid Duplexes

Abstract

DNA:RNA hybrid duplex plays important roles in various biological processes. Both its structural stability and interactions with proteins are highly sequence dependent. In this study, we utilize homebuilt optical tweezers to investigate how GC contents in the sequence influence the structural and mechanical properties of DNA:RNA hybrid by measuring its contour length, elasticities, and overstretching dynamics. Our results support that the DNA:RNA hybrid adopts a conformation between the A-and B-form helix, and the GC content does not affect its structural and elastic parameters obviously when varying from 40 to 60% before the overstretching transition of DNA:RNA hybrid occurs. In the overstretching transition, however, our study unravels significant heterogeneity and strong sequence dependence, suggesting the presence of a highly dynamic competition between the two processes, namely the S-form duplex formation (nonhysteretic) and the unpeeling (hysteretic). Analyzing the components left in DNA:RNA hybrid after the overstretching transition suggests that the RNA strand is more easily unpeeled than the DNA strand, whereas an increase in the GC content from 40 to 60% can significantly reduce unpeeling. Large hysteresis is observed between the stretching and relaxation processes, which is also quantitatively correlated with the percentage of unpeeling in the DNA:RNA duplex. Increasing in both the salt concentration and GC content can effectively reduce the hysteresis with the latter being more significant. Together, our study reveals that the mechanical properties of DNA:RNA hybrid duplexes are significantly different from double-stranded DNA and double-stranded RNA, and its overstretching behavior is highly sequence dependent. These results should be taken into account in the future studies on DNA:RNA-hybrid-related functional structures and motor proteins.

Stretching of Long Double-Stranded DNA and RNA Described by the Same Approach

“…We show that useful information can be easily extracted and some nontrivial trends explained without the need to perform MD simulations. In what follows, we apply the algorithm (Figure , Section ) to experimental data for dsDNA with 48% and 70% GC content (Figure ), dsDNA with 0% GC content (poly­(dA-dT)) and dsDNA with 100% GC content (poly­(dG-dC)) (Figure ), and a DNA/RNA hybrid duplex with 52% GC content (Figure ). DNA/RNA hybrid duplexes , occur in various biological processes − and have potential in biomedical applications, including gene silencing.…”

“…In what follows, we apply the algorithm (Figure , Section ) to experimental data for dsDNA with 48% and 70% GC content (Figure ), dsDNA with 0% GC content (poly­(dA-dT)) and dsDNA with 100% GC content (poly­(dG-dC)) (Figure ), and a DNA/RNA hybrid duplex with 52% GC content (Figure ). DNA/RNA hybrid duplexes , occur in various biological processes − and have potential in biomedical applications, including gene silencing.…”

“…The blue line, F ( L x / L 0 ), is the model force–extension curve of the polymer at its minimum energy state. The parameters of the model and the eWLC (Table 1 from ref ) are shown in the figure. The helical rise of 3.03 Å is assumed .…”

Folding Free Energy Determination of an RNA Three-Way Junction Using Fluctuation Theorems

“…Briefly, the elastic response of the ssRNA and hybrid DNA-RNA handles is calculated using the inextensible worm-like chain model [ 36 ]. The elastic parameters ( p for persistence length and for the interphosphate distance) used in this study are: nm and nm/base for the ssRNA and nm and nm/basepair for the hybrid handles [ 35 , 37 , 38 , 39 , 40 , 41 ]. The folded hairpin is modeled as a freely-rotating dipole of 2 nm length [ 37 ].…”

Single-molecule mechanical unfolding kinetics of unmodified Saccharomyces cerevisiae tRNAPhe: a hint to the tRNA chaperone-tRNA interaction mechanism

“…Single-molecule pulling experiments were performed using home-built single-trap optical tweezers described previously (30). Briefly, a linearly polarized TEM 00 -mode 1064-nm laser beam (Changchun New Industries Optoelectronics Technology, Changchun, China) was coupled to a polarization-maintaining single-mode fiber (OZ Optics, Ottawa, Canada).…”