Mechanical Properties of Nucleic Acids in Bulk and Under Confinement and Their Interaction with Peptide

Abstract

Proteins, DNA and RNA are arguably the most studied biological molecules with curiously interconnected relation that appears ubiquitous in all organisms. Almost all nucleoprotein interactions and DNA manipulation events involve mechanical deformations of DNA. The flexibility of RNA and DNA is crucial for their biological functions. Apart from DNA and RNA, peptide nucleic acids (PNAs), although synthetic nucleic acids, are being widely studied due to their potential application in different fields of biotechniques. PNAs hybridize with complementary DNAs or RNAs with extraordinary affinity, exhibiting remarkable thermal stability.

During interaction with different biomolecules like protein, the nucleic acids modulate their conformations by bending, stretching and twisting such that the shape and size modifications optimize the Watson-Crick geometry for effective interaction. The study of the elasticity of the nucleic acid duplexes is therefore critical. In this thesis, using all-atom molecular dynamics (MD) simulations, we have studied the structure, mechanics and thermodynamics of double-stranded (ds) DNA, RNA, PNA and their hybrid duplexes. We have computed bending persistence length, stretch modulus and torsional stiffness of the duplexes along with different structural and helicoidal parameters. We have also computed the effect on these properties for dsDNA with oxidative modification of the bases and with confinement inside the single-walled carbon nanotube (SWCNT). In addition, we have analysed the interaction affinity of the dsDNA with cationic peptide (mimicking protamine) in both native and charge modified (phosphorylated) states of the peptide.

We found dsPNA to have higher bending, stretching and twisting flexibility than dsDNA and dsRNA, indicating that dsPNA may easily change its structure to make complexes with biomolecules. The PNA-DNA and PNA-RNA hybrid duplexes have elastic properties between dsPNA and dsDNA/dsRNA. We argue that the neutral backbones of the PNA make it less stiff than dsDNA and dsRNA molecules. We found that the oxidative modification of dsDNA does not weaken the Watson-Crick geometry. Instead, it is strengthened with the increase in electrostatic interaction. The oxidative damage changes the mechanical, helical and groove parameters of dsDNA, and these changes are more significant on twisting and stretching flexibilities compared to bending stiffness. With oxidation, the bending flexibility of dsDNA gets suppressed, and the oxidized dsDNAs are more flexible to stretch and twist than the non-oxidized dsDNA. Compared to the global mechanical properties, the changes in helical and groove properties are found to be more prominent, concentrated locally at the oxidation sites, and causing the transition of the backbone conformations from BI to BII at the regions of oxidative damage.

We observed the denaturation of dsDNA when confined inside narrower SWCNT, and the denaturation is diameter dependent, with a critical diameter of 3.25 nm. Below this critical diameter of SWCNT, the dsDNA is denatured, and above it, the dsDNA retains its native structure. The elastic properties of the dsDNA in cylindrical confinement inside SWCNTs are also found to be diameter dependent. Inside moderately wide SWCNTs, the confined dsDNA is highly rigid to bend, stretch and twist, whereas, inside wider SWCNTs, the elastic properties change surprisingly, making the dsDNA highly flexible.

Regarding the interaction of dsDNA with protamine-like cationic peptide, we found that phosphorylation of the residues of cationic peptides decreases their binding affinity with dsDNA. Among the two events, phosphorylation and dephosphorylation of the protamines, phosphorylation minimizes the shapes of protamines (making them more spherical), reducing the interaction with the nucleobases of dsDNA, and results in weak binding. Also, the level of compaction of dsDNAs under the influence of the phosphorylated peptides is found to be weaker than that caused by non-phosphorylated peptides. Because of the high binding affinity of dephosphorylated (non-phosphorylated) cationic peptides, the dephosphorylation of peptides is responsible for making dsDNA duplexes more compacted.

Thus, the works carried out here in this dissertation explain the different mechanical and thermodynamic properties of the various nucleic acids (namely; dsDNA, dsRNA, dsPNA and their hybrids), the structure and thermodynamic stability of dsDNA inside the confined space of SWCNT, and the effect of phosphorylation of peptides in the DNA compaction process.