J. Chakrabarti

Dipolar Solute Rotation in a Supercritical Polar Fluid

Fluorescence anisotropy measurements reveal a non-monotonic density dependence for average rotation time (τ(R)) of a polar solute, coumarin153 (C153) in polar supercritical fluoroform (CHF(3)). The conventional Stokes-Einstein-Debye model, relating τ(R) to the solvent viscosity, fails to explain the observed density dependence, because the experimental viscosity increases monotonously with density for a fluid, in general. Here, the density-dependent τ(R) is calculated by incorporating the wave vector-dependent viscosity of the solvent and the solute-solvent interaction. A molecular hydrodynamic description is used for the wave vector-dependent viscosity which is verified by molecular dynamics (MD) simulation. A justification for the applicability of the present prescription is provided by reproducing the experimental viscosity of supercritical (SC) CHF(3). Solute-solvent interaction has been included via the fluctuating torque acting on the rotating solute. Incorporation of wave vector-dependent viscosity leads to a qualitative description of the experimental density dependence of τ(R) which is further improved upon inclusion of solute-solvent interaction.

Structural properties of polymeric DNA from molecular dynamics simulations

Most of the reported DNA structural studies are based on oligonucleotide structures, which have artifacts due to unstable terminal base pairs (bps). We have carried out molecular dynamics simulation of DNA oligonucleotides in such a manner that gives rise to properties of polymeric DNA of infinite length. Molecular dynamics simulation studies of six homo- and heteropolymeric DNA sequences are reported here to understand structural features of all ten unique dinucleotide sequences. We observe that each of these dinucleotide sequences has unique features in agreement with Calladines rule [C. R. Calladine, J. Mol. Biol. 161, 343 (1982)]. We noticed significant structural alternation between B(I) and B(II) forms for d(CA).d(TG) dinucleotide, where one of the strands showed frequent transitions between usual and unusual epsilon and zeta torsion angles associated with bp stacking geometry. In terms of the calculated bending rigidity and persistence length, pyrimidine-purine bp steps, namely, d(TA).d(TA), d(CA).d(TG), and d(CG).d(CG) are the most flexible dinucleotide bp steps. We estimated the major groove widths from our simulations. We did not observe much variation in major and minor groove widths depending on the base sequence. However, the distribution of water molecules in the minor groove shows sensitivity to the DNA sequence.

Changes in Thermodynamic Properties of DNA Base Pairs in Protein-DNA Recognition

Abstract The mechanism of protein-DNA recognition, particularly the induced fit mechanism, is poorly understood due to ineffective analysis of the protein-DNA complex crystal structures. It is expected that upon protein binding the DNA becomes structurally more rigid. However, a previous analysis (W.K. Olson, A. A. Gorin, X. Lu, L. M. Hock and V. Zhurkin, Proc. Natl. Acad. Sci. USA, 95, 11163 (1998)) indicates increase in the flexibility of the DNA segment complexed with protein. We have considered an ensemble of configurations from crystallographic data of the TBP-TATA box complex structures under a given thermodynamic condition. Analysis of the ensemble of structures of this complex indicates that the DNA deforms significantly to form specific hydrogen bonds and as a consequence, its structure attains more rigidity. We calculate the free energy profiles in term of the DNA base pair (bp) step parameters via the binding patterns in the ensemble of the given complex, and for free DNA bp steps as well. The rigidities estimated from these free energies for small deformations around the minimum indicate enhanced structural rigidities of DNA upon complexation with protein. Further, the changes in the thermodynamic properties of the bp steps upon complex formation have been estimated from the two sets of free energy profiles. These results indicate differential role played by different bp steps in the thermodynamic stabilization of the complex.

Solute rotation in polar liquids: Microscopic basis for the Stokes-Einstein-Debye model

Here, we develop a framework for a molecular level understanding of the celebrated Stokes-Einstein-Debye (SED) formula. In particular, we explore reasons behind the surprising success of the SED model in describing dipolar solute rotation in complex polar media. Relative importance of solvent viscosity and solute-solvent dipolar interaction is quantified via a self-consistent treatment for the total friction on a rotating solute where the hydrodynamic contribution is modified by the friction arising from the longer ranged solute-solvent dipolar interaction. Although the solute-solvent dipolar coupling is obtained via the Mori-Zwanzig formalism, the inclusion of solvent structure via the wave vector dependent viscosity in the hydrodynamic contribution incorporates solvent molecularity in the present theory. This approach satisfactorily describes the experimental rotation times measured using a dipolar solute, coumarin 153 (C153), in protic and aprotic polar liquids, and more importantly, provides microscopic explanation for insignificant contribution of electrical interactions on solute rotation, in contrast to the substantial role played by the translational dielectric friction in the context of ionic mobility. It is also discussed on how the present theory can be suitably extended to study the rotation of a realistic solute in media other than dipolar solvents.

Microscopic mechanisms of confinement-induced slow solvation.

Several studies show that the dynamics of solvent molecules around a solute slows down in a nanoscale confined geometry compared to the bulk condition. Here we perform numerical simulations to investigate the microscopic mechanisms of such slowing down. We show a substantial slowing down of solvation dynamics around a solute in strong solvophilic confinements due to suppression of fluid diffusion in the presence of solvophilic walls, along with restricted solvent dynamics due to geometrical constraints. The solvation in strong solvophobic confinements becomes slower than the same in the bulk as well, but not as significantly as in the solvophilic case. This is due to the competition between restriction in solvent dynamics and faster in-plane solvent diffusion. We place our findings in perspective of various solvation dominated chemical processes in nanoconfined geometry.

Reversible thermal unfolding of a yfdX protein with chaperone-like activity

yfdX proteins are ubiquitously present in a large number of virulent bacteria. A member of this family of protein in E. coli is known to be up-regulated by the multidrug response regulator. Their abundance in such bacteria suggests some important yet unidentified functional role of this protein. Here, we study the thermal response and stability of yfdX protein STY3178 from Salmonella Typhi using circular dichroism, steady state fluorescence, dynamic light scattering and nuclear magnetic resonance experiments. We observe the protein to be stable up to a temperature of 45 °C. It folds back to the native conformation from unfolded state at temperature as high as 80 °C. The kinetic measurements of unfolding and refolding show Arrhenius behavior where the refolding involves less activation energy barrier than that of unfolding. We propose a homology model to understand the stability of the protein. Our molecular dynamic simulation studies on this model structure at high temperature show that the structure of this protein is quite stable. Finally, we report a possible functional role of this protein as a chaperone, capable of preventing DTT induced aggregation of insulin. Our studies will have broader implication in understanding the role of yfdX proteins in bacterial function and virulence.

Quantum Chemical Studies on Stability and Chemical Activities in Calcium Ion Bound Calmodulin Loops

Quantum chemical (QC) calculations for macromolecules require truncation of the molecule, highlighting the portion of interest due to heavy computation cost. As a result, an estimation of the effects of truncation is important to interpret the energy spectrum of such calculations. We perform density functional theory based QC calculations on calcium ion bound EF-hand loops of Calmodulin isolated from the crystal structure in an implicit solvent. We find that the terminal contributions of neutral capping are negligible across the entire ground-state energy spectrum. The coordination energy range and the nature of hybridization of the coordination state molecular orbitals remain qualitatively similar across these loops. While the HOMO and LUMO of loops in the N-terminal domain are dominated by the acidic aspartates, and the polar/hydrophobic residues, respectively, these levels of the C-terminal domain loops show strong localized electron density on the phenyl rings of the tyrosines. The Fukui index calculation identifies the hydroxyl oxygen in the phenyl ring of Y99 as a potent nucleophile. Our analysis indicates a general way of interpreting the electronic energy spectra to understand stability and functions of large biomolecules where the truncation of the molecule and, hence, the terminal capping effects are inevitable.

SDS induced dissociation of STY3178 oligomer: experimental and molecular dynamics studies

STY3178 is a yfdX protein from Salmonella Typhi. yfdX proteins occur ubiquitously in a number of virulent bacteria but their cellular localization is unknown. Our earlier studies have shown that STY3178 is a trimer and can be a periplasmic chaperone protein. In the present study we show the stability of STY3178 in the presence of the bio-mimetic anionic surfactant sodium dodecyl sulphate (SDS). With increasing concentrations of SDS we observe monomeric STY3178 which reversibly forms the trimer upon decreasing the surfactant concentration. Protein tertiary structure is not perturbed in the presence of SDS. We show using molecular dynamics simulation and conformational thermodynamics data that SDS induces stability of the monomer compared to an isolated monomer of STY3178. This supports our experimental observations.

Molecular dynamics studies on conformational thermodynamics of Orai1–calmodulin complex

Molecular understanding of bio-macromolecular binding is a challenging task due to large sizes of the molecules and presence of variety of interactions. Here, we study the molecular mechanism of calmodulin (CaM) binding to Orai1 that regulates Ca2+-dependent inactivation process in eukaryotic cells. Although experimental observations indicate that Orai1 binds to the C-terminal of Ca2+-loaded CaM, it is not decisive if N-domain of CaM interacts with Orai1. We address the issue of interaction of different domains of CaM with Orai1 using conformational thermodynamic changes, computed from histograms of dihedral angles over simulated trajectories of CaM, CaM-binding domain of Orai1 and complexes of CaM with Orai1. The changes for all residues of both C and N terminal domains of CaM upon Orai1 binding are compared. Our analysis shows that Orai1binds to both C-terminal and N-terminal domains of CaM, indicating 1:2 stoichiometry. The Orai1 binding to N-terminal domain of CaM is less stable than that to the C-terminal domain. The binding residues are primarily hydrophobic. These observations are in qualitative agreement to the experiments. The conformational thermodynamic changes thus provide a useful computational tool to provide atomic details of interactions in bio-macromolecular binding.

Response to chemical induced changes and their implication in yfdX proteins

yfdX proteins occur in a large number of virulent bacteria. Recently we have shown that STY3178, a yfdX protein from Salmonella Typhi, exists in a trimeric state in solution which is capable of interacting with antibiotics, stable at elevated temperatures and undergoes reversible thermal unfolding. In this present study, we report the chemical response of STY3178. We monitor the stability of the protein in presence of chaotropes. It can regain the native-like structure from the chaotrope induced unfolded states. The structural stability of this protein is further studied in a wide pH range which reveals that the STY3178 trimer is stable in both acidic as well as basic media. We further show that the protein interacts with oxalate in vitro. Finally, we perform computational studies viz. modeling and molecular dynamics simulation to understand the stability of trimeric STY3178 over its monomer conformation. The conformational thermodynamic changes indicate that oligomerization induces stability via salt bridge interactions, present at the monomer interface.