Synthesis, characterization and multi-spectroscopic DNA/HSA interaction studies of synthetic human Follicle-Stimulating Hormone Beta 33-53 peptide conjugated PEGylated graphene oxide nanoparticles.

The objective of the current study was to synthesize, characterize and explore the interaction of PEGylated graphene oxide (pGO) and synthetic human Follicular stimulating hormone β 33-53 peptide conjugated PEGylated graphene oxide nanoparticles (pGO-FSH) with human serum albumin (HSA) and calf thymus DNA (CT-DNA). The pGO/ pGO-FSH nanoparticles were synthesized using a modified Hummer's method, and the FSH peptide was conjugated through a maleimide crosslinking reaction. Synthesized nanoparticles were then characterized using techniques like FT-IR, UV-Visible absorbance, CD and Raman spectroscopy, and XRD and TGA. Morphological and particle size analysis was studied using SEM, TEM, DLS, and zeta potential measurements. The presence of FSH β 33-53 peptide was confirmed qualitatively and quantitatively using CD spectroscopy and Bradford's assay. Binding studies of pGO/pGO-FSH nanoparticles with HSA and DNA were carried out using biophysical techniques. The complex formation between pGO/pGO-FSH nanoparticles and HSA was revealed by UV absorbance spectroscopy, and the observed fluorescence quenching was confirmed by steady-state fluorescence spectroscopy. Time-resolved fluorescence quenching studies have shown that dynamic quenching plays an important role in binding HSA with pGO/pGO-FSH nanoparticles. However, structurally no significant changes were observed in the native structure of HSA upon binding with pGO/pGO-FSH nanoparticles suggesting that the latter did not induce any structural distortions together, confirmed by DSC, FT-IR, and CD spectroscopy experimental findings. Binding constants and thermodynamic parameters calculated using double logarithmic and Van't Hoff plots suggested weak and moderate binding affinity along with the involvement of hydrophobic and hydrogen bonding interactions between HSA and pGO/pGO-FSH nanoparticles, respectively. UV absorbance and fluorescence spectroscopy have revealed that pGO/pGO-FSH nanoparticles interact with DNA by binding at the minor groove region. These findings were further confirmed by DNA melting and viscosity studies. CD and FT-IR spectroscopy studies have shown no changes in the helical structure of B-form of DNA, thereby emphasizing the groove-binding nature of pGO/pGO-FSH nanoparticles. The obtained results are useful in further considering the potentiality of pGO-FSH nanoparticles as drug-delivery systems for in vivo applications, especially to target ovarian cancer.