Abstract:
Biomolecular machines are responsible for carrying out a host of essential cellular processes.In accordance to the wide range of functions they execute, the architectures of these also vary greatly. Yet, despite this diversity in both structure and function, they have some common characteristics. They are all large macromolecular complexes that enact multiple steps during the course of their functions. They are also 'Brownian' in nature, i.e., they rectify the thermal motions of their surroundings into work. Yet how these machines can utilise their surrounding thermal energy in a directional manner, and do so in a cycle over and over again, is still not well understood. My research during my Ph.D. has spanned the development, evaluation and use of biophysical, in particular single-molecule, tools in the study of the functional mechanisms of biomolecular machines, particularly, the bacterial ribosome. In this talk, I will describe how a mathematical framework we developed to relate any experimental data to an ideal template irrespective of the scale, background and noise, may be used to evaluate the extent of spatial information in cryogenic electron microscopy (cryoEM) density maps. This application will not only aid the study of biomolecular structure using cryoEM by structural biologists, but also enable biophysicists and biochemists who use structural models to interpret and design their experiments to evaluate the cryoEM data they need to use for their investigations. Additionally, I will describe the use of temperature-dependent single-molecule kinetic experiments to elucidate a mechanism by which ligands can modulate and drive the conformational dynamics of the ribosome in a manner that facilitates ribosome-catalysed protein synthesis. This mechanism has implications to our understanding of the functional mechanisms of the ribosome in particular, and of biomolecular machines in general.
