Study of Biomolecular Recognition Using Time-Resolved Optical Spectroscopy

Molecular recognition process refers to the weak non-covalent interaction, which takes place selectively and specifically between small ligand molecules with biological macromolecules. Understanding of such recognition in biological and biomimetic milieu is the central attraction for drug designing, which is crucial for the improvement of human healthcare. A thorough knowledge of the structural, dynamical and energetic parameters that dictate such molecular interactions can find immense use in the modulations of the ligand-macromolecule recognition process. In this article, we present our continuous effort to investigate the fundamental physical processes involved in the biomolecular recognition, e.g. efficiency (binding affinity and rigidity of the complex) and role of solvent molecules in the molecular recognition using steady state and predominantly, ultrafast time-resolved fluorescence spectroscopy. In this perspective, we have thoroughly investigated the molecular recognition of small ligand/drug molecules (Rifampicin; Rf, 4-(dicyanomethylene)-2-methyl-6-(p-dimethylaminostyryl)-4H-pyran; DCM, and Nile Blue; NB) by a human transporter protein, Human Serum Albumin (HSA), and also established the nonspecific type of interaction between a ligand molecule (Rf) and a biomimetic system (Sodium Dodecyl Sulfate (SDS) micelle). Simultaneous recognition of an intercalator (Ethidium Bromide, EtBr) and a DNA minor groove binder (Hoeschst 33258, H258) to a dodecamer DNA of specific sequence has also been monitored. Besides, we report an investigation on the recognition of ethidium (Et) cation, a potential mutagen, by synthetic DNA and various cell nuclei in presence of a stimulant drug, caffeine, employing the mentioned spectroscopic techniques along with NMR and fluorescence microscopy. Moreover, we have explored the differential molecular recognition of 8-anilino-1-naphthalenesulfonic acid (ANS) and 2,6-p-toluidinonaphthalene sulfonate (TNS) by bovine pancreatic -chymotrypsin (CHT) upon interaction with genomic DNA. The correlation of the molecular recognition of the DNA and DNA-protein complexes with the hydration dynamics has been further exploited in our studies. In addition, we have developed functional nanoparticles/Quantum dots (QDs) that are covalently linked to biological molecules to detect the molecular interaction phenomenon between biomolecules. It should be noted that QDs have a significant contribution in the field of nano-biotechnology due to its high quantum yield, low photo-bleaching and increased biological application (cell labeling, in vivo imaging, gene delivery, sensing of fluorescence and molecular recognition). In this regard, we have exploited QDs as a potential energy donor/acceptor system and validated Forster resonance energy transfer (FRET) model over nano-surface energy transfer (NSET) technique. Therefore, the ultrafast non-radiative energy migration from tryptophan (Trp214) present in HSA to the HSA bound CdS QD, and from 4-nitrophenyl anthranilate (NPA) to Silver (Ag) nanoclusters in CHT (both NPA and Ag bound to CHT) have been investigated using FRET technique to monitor the protein folding pathway of HSA, and molecular interaction between NPA and CHT respectively. Moreover, we have also used functionalized QDs (CdSe/ZnS) for the detection of molecular recognition of ethidium bromide (EtBr) by a synthetic DNA. However, the intention of this review is to give an overview of ultrafast optical spectroscopic techniques for the exploration of biomolecular recognition, which may find potential significance for further research in the field of nano-biotechnology and medicine