Research | Deepak Chatterjee

RESEARCH INTEREST

Integrin Reuglation-Analyses of Multi protein ensembles of the integrins cytosolic tails

Integrins are crucial transmembrane heterodimeric receptors involved in cell adhesion, migration, proliferation and organismal development. The subfamily of leukocyte-restricted ß2 integrins consists of four members (αLß2, αMß2, αXß2 and αDß2). They play essential roles in immune responses, including leukocyte trafficking into tissues, phagocytosis, antigen-presentation, and immune tolerance. Like other members of the integrin family, the ß2 integrins have relatively short cytoplasmic domains (CDs). These CDs serves as important binding sites for an expanding list of cytoplasmic molecules that either regulate integrin ligand-binding or integrin-mediated cellular signalling.

To date, a number of positive regulators of integrin functions have been identified, including cytoplasmic molecules talin and 14-3-3ζ. On the other hand, negative regulators of integrins such as Dok1 and filamin have been reported. Many of these regulators have overlapping binding sites on the integrin CDs. Whether these molecules are mutually exclusive and how they are spatio-temporally regulated with respect to integrin activation remains to be elucidated. Although, regulatory proteins interact with the CDs, however, it remains unclear whether direct interactions occur between positive regulators and negative regulators. The aim of my research to characterize these intermolecular associations so as to provide mechanistic insights into ß2 integrins signalling and regulation. The specific aims are as follow.

1. To characterize direct interactions between the positive regulator 14-3-3ζ with negative regulators Dok1 and filamin A, and their effects on integrin ß2 CD interaction.
2. To determine the structures and interactions of IgFLN21 domain of filamin A in complex with integrin ß2 and αM or αL CDs.
3. To determine a quaternary complex of IgFLN21 domain of filamin A with phosphorylated ß2 with 14-3-3ζ and Talin F3 domain.
4. To determine the role of phosphorylation in ß2 and ß3 CD for Integrin.
5. Investigating the mechanism involve in Kindlin as co-activator with Talin/ ß1 rather than direct activator

My research is of academic significance because to our knowledge, there is limited information on the aforementioned intermolecular interactions. Further, it will provide novel insights into the molecular basis of overt leukocyte responses under pathological conditions, such as systemic lupus erythematosus. Data from these studies are valuable for development anti-inflammation therapeutics

Insulin and its analogs-Structure, interaction and Dynamics

We have exploited various insulin analog to elucidate the underlying structure and dynamics of insulin as a monomer in solution and aligned media. A model was provided by insulin lispro (the active component of Humalog®; Eli Lilly and Co.). Whereas NMR-based modeling recapitulated structural relationships of insulin crystals (T-state protomers), dynamic anomalies were revealed by amide-proton exchange kinetics in D2O. Surprisingly, the majority of hydrogen bonds observed in crystal structures are only transiently maintained in solution, including key T-state-specific inter-chain contacts. Long-lived hydrogen bonds (as defined by global exchange kinetics) exist only at a subset of four α-helical sites (two per chain) flanking an internal disulfide bridge (cystine A20–B19); these sites map within the proposed folding nucleus of proinsulin. The anomalous flexibility of insulin otherwise spans its active surface and may facilitate receptor binding. Because conformational fluctuations promote the degradation of pharmaceutical formulations, we envisage that “dynamic re-engineering” of insulin may enable design of ultra-stable formulations for humanitarian use in the developing world.

Structure and Dynamics of peptides in model membrane

The lethal Coronaviruses (CoVs), Severe Acute Respiratory Syndrome-associated Coronavirus (SARS-CoV) and most recently Middle East Respiratory Syndrome Coronavirus, (MERS-CoV) are serious human health hazard. A successful viral infection requires fusion between virus and host cells carried out by the surface spike glyco-protein or S protein of CoV. Current models propose that the S2 subunit of S protein assembled into a hexameric helical bundle exposing hydrophobic fusogenic peptides or fusion peptides (FPs) for membrane insertion. The N-terminus of S2 subunit of SARS-CoV reported to be active in cell fusion whereby FPs have been identified. Atomic-resolution structure of FPs derived either in model membranes or in membrane mimic environment would glean insights toward viral cell fusion mechanism. Here, we have solved 3D structure, dynamics and micelle localization of a 64-residue long fusion peptide or LFP in DPC detergent micelles by NMR methods.