Our research programme aims to innovate and engineer novel technologies to help understand and manage infectious diseases using single molecule detection, single cell analysis, quantitative genomics and high-resolution imaging. Using highly sensitive detection of single biological macromolecules like proteins or nucleic acids by both optical and non-optical methods, we determine the molecular behaviour at the single molecule level as well as its distribution at the population level.
TOOLS FOR PROBING AND QUANTIFYING BIOMOLECULES AT THE SINGLE CELL AND SINGLE MOLECULE LEVEL
We have been extending single molecule based imaging and spectroscopic techniques like single molecule Förster resonance energy transfer (FRET), single molecule protein induced fluorescence enhancement (PIFE), single particle tracking and super-resolution microscopy that allow quantification of cellular composition even for low copy number constituents, rapid live cell imaging with high-resolution spatial localization. As part of the larger goal to understand cellular heterogeneity, we have developed a high throughput Lab-on-Chip device to quantify absolute copy number of nucleic acids in single cells.
UNDERSTANDING PRINCIPLES OF MACROMOLECULAR SELF-ASSEMBLY
Another key research focus is in the area of understanding self-assembly of bio-macromolecular structures, we have been pursuing studies of 2D assembly of bacterial pore forming toxins and 3D assembly of viruses/bacteriophages. For pore-forming toxins from bacteria, we find the first set of conformational changes that ensure protein insertion into bilayer membranes is enhanced by the presence of cholesterol. Cholesterol is a key mammalian cell membrane constituent that is absent in bacterial membranes can explain how these proteins selectively puncture across mammalian cell membranes. Separately, we monitor bacteriophage assembly on live bacteria using super-resolution microscopy allowing us to probe key steps in the assembly of bacteriophages that are technologically relevant.
PROBING VIRUSES USING QUANTITATIVE SINGLE VIRUS SEQUENCING AND MODELING
RNA viruses propagate as a quasispecies of closely related genotypes in the host. This helps the virus in rapid adaptation, acquiring drug resistance and fast expansion. Sequence diversity and virulence of a viral species is regulated by the host selection pressures at different levels of its life cycle. Our main objective here has been to define such genetic contributions of the full-length virus genome to its infectivity, which is term as “infectivity fitness”. To probe the sequence-infection relationship, we have developed a method that would give us the sequence of each individual viral RNA molecules along with quantitative copy numbers and thereby providing the tools to track the sequence variations in a viral population. Combined with data at various stages of virus infection and modeling of the viral RNA as a substrate for the various viral processes allows us to quantify and model virus dynamics during its lifecycle.