Proteins and nucleic acids are ubiquitous biopolymers, and, to function, they are required to undergo specific conformational transitions spanning over different time regimes and distances. These include global folding dynamics to a thermodynamically favorable state and organized local structural motions, as well as spontaneous conformational fluctuations. Therefore, gaining a molecular-level understanding of the structure-dynamics-function relationship of any biopolymer requires detailed information about its conformational dynamics landscape. However, yielding such information via spectroscopic techniques has often been challenging due to the fact that conventionally acquired spectroscopic signals often lack the desired spatial and temporal resolution. To overcome this limitation, various strategies and methods have been explored. In this thesis, we focus on the development and utility of various chemical moieties, such as molecular cross-linkers and unnatural amino acids, as site-specific conformational constraints, and spectroscopic probes. The former provides a means to reveal ‘hidden’ conformational events and dynamics, whereas the latter allows detection of specific molecular interactions and structural transitions. Specifically, using this chemical modification approach and multiple spectroscopic methods, we have determined 1) the nucleation site and rate of α-helix folding, 2) how ionic liquids interact with and destabilize a multi-domain copper binding protein, Azurin, (3) the protein denaturing effect of a naturally occurring stabilizing osmolyte, glycine betaine, and (4) the effect of Hofmeister anions on the hydrogen-bonding strength of water. In addition, we have demonstrated that the blue fluorescent amino acid, 4-cyanotryptophan, is a useful protein hydration reporter and that 4-cyanoindole-2′-deoxyribonucleoside can serve as a site-specific dual fluorescence and infrared probe of DNA structure and dynamics.