Kalyan Sundar Ghosh

Studies on the interaction of copper complexes of (-)-epicatechin gallate and (-)-epigallocatechin gallate with calf thymus DNA.

The interaction of copper complexes of (-)-epicatechin gallate (ECG) and (-)-epigallocatechin gallate (EGCG) with calf thymus DNA (ct-DNA) was investigated by UV-visible (UV-Vis), fluorescence and circular dichroism along with melting studies. It was observed that both copper complexes quench the fluorescence intensity of ethidium bromide bound ct-DNA upon binding, resulting in a ground state complex formation by a static quenching process. The binding constants evaluated from fluorescence data were supported by the UV-Vis study. The values ranged from 0.84 to 1.07x10(5)M(-1) and 1.14 to 1.04x10(5)M(-1) for Cu(II)-ECG and Cu(II)-EGCG, respectively for the temperature range 21-42 degrees C with two binding sites. Thermodynamic parameters obtained are suggestive of the involvement of different modes of interaction during binding for each complex although both were found to be intercalating in nature. Circular dichroism studies and variations in the melting temperature reveal unwinding of the ct-DNA helix with conformational changes due to binding.

A spectroscopic investigation into the interactions of 3′-O-carboxy esters of thymidine with bovine serum albumin

Binding studies of 3′‐O‐carboxy esters of thymidine, reported inhibitors of ribonucleases, with bovine serum albumin (BSA) have been explored in this report. Fluorescence spectroscopy in combination with Fourier transform infrared (FTIR) and circular dichroism (CD) spectroscopy have been used to determine the nature and mode of binding. The binding and quenching parameters were determined from tryptophan fluorescence quenching by Scatchard plots and modified Stern–Volmer plots. The association constants are of the order of 104 M−1 for both the ligands. Thermodynamic parameters suggest that apart from an initial hydrophobic association, hydrogen bonding and van der Waals interactions play a decisive role during protein‐ligand complex formation. Minor changes were observed in the secondary structures of human serum albumin (HSA) as revealed by FTIR and CD. Docking studies suggest that the ligands are close to Trp 213, which causes fluorescence quenching.

Evidence of conformational changes in adsorbed lysozyme molecule on silver colloids

In this article, we discuss metal-protein interactions in the Ag-lysozyme complex by spectroscopic measurements. The analysis of the variation in relative intensities of SERS bands reveals the orientation and the change in conformation of the protein molecules on the Ag surface with time. The interaction kinetics of metal-protein complexes has been analyzed over a period of 3h via Raman measurements. Our analysis indicates that the Ag nanoparticles most likely interact with Trp123 which is in close proximity to Phe34 of the lysozyme molecule.

Copper complexes of ( )-epicatechin gallate and ( )-epigallocatechin gallate act as inhibitors of Ribonuclease A

Green tea polyphenols, which have the ability to inhibit angiogenesis, form complexes with Cu(II), a known potent stimulator of blood vessel proliferation. Copper complexes of (−)‐epicatechin gallate and (−)‐epigallocatechin gallate were found to inhibit the enzymatic activity of Ribonuclease A (RNase A) as revealed by an agarose gel based assay and urea denatured gel electrophoresis. The copper complexes were found to be non‐competitive inhibitors of RNase A with inhibition constants in the micromolar range. Changes in the secondary structure of the protein are found to occur due to the interaction as revealed from Fourier transform infrared and circular dichroism studies.

An investigation into the altered binding mode of green tea polyphenols with human serum albumin on complexation with copper.

Green tea is rich in several polyphenols, such as (−)-epicatechin-3-gallate (ECG), (−)-epigallocatechin (EGC), and (−)-epigallocatechin-3-gallate (EGCG). The biological importance of these polyphenols led us to study the major polyphenol EGCG with human serum albumin (HSA) in an earlier study. In this report, we have compared the binding of ECG, EGC, and EGCG and the Cu(II) complexes of EGCG and ECG with HSA. We observe that the gallate moiety of the polyphenols plays a crucial role in determining the mode of interaction with HSA. The binding constants obtained for the different systems are 5.86 ± 0.72 × 104 M−1 (K ECG-HSA), 4.22 ± 0.15 × 104 M−1 (K ECG-Cu(II)-HSA), and 9.51 ± 0.31 × 104 M−1 (K EGCG-Cu(II)-HSA) at 293 K. Thermodynamic parameters thus obtained suggest that apart from an initial hydrophobic association, van der Waals interactions and hydrogen bonding are the major interactions which held together the polyphenols and HSA. However, thermodynamic parameters obtained from the interactions of the copper complexes with HSA are indicative of the involvement of the hydrophobic forces. Circular dichroism and the Fourier transform infrared spectroscopic measurements reveal changes in α-helical content of HSA after binding with the ligands. Data obtained by fluorescence spectroscopy, displacement experiments along with the docking studies suggested that the ligands bind to the residues located in site 1 (subdomains IIA), whereas EGC, that lacks the gallate moiety, binds to the other hydrophobic site 2 (subdomain IIIA) of the protein.

Inhibition of Ribonuclease A by polyphenols present in green tea

We report the effect of the natural polyphenolic compounds from green tea on the catalytic activity of Ribonuclease A (RNase A). The compounds behave as noncompetitive inhibitors of the protein with inhibition constants ranging from 80–1300 μM. The dissociation constants range from 50–150 μM for the RNase A–polyphenol complexes as determined by ultraviolet (UV) and circular dichroism (CD) studies. We have also investigated the changes in the secondary structure of RNase A on complex formation by CD and Fourier transformed infrared (FTIR) spectroscopy. The presence of the gallate moiety has been shown to be important for the inhibition of enzymatic activity. Docking studies for these compounds indicate that the preferred site of binding is the region encompassing residues 34–39 with possible hydrogen bonding with Lys 7 and Arg 10. Finally we have also looked at changes in the accessible surface area of the interacting residues on complex formation for an insight into the residues involved in the interaction. Proteins 2007.

Characterization of the structure of the phosphoprotein of Chandipura virus, a negative stranded RNA virus probing intratryptophan energy transfer using single and double tryptophan mutants

The phosphoprotein (P protein) of Chandipura virus (CHPV), a negative stranded RNA virus, is involved in both transcription and replication phases of the viral life cycle. The two Tryptophan (Trp) residues of CHPV, located at 105 and 135 respectively and two single Trp mutants W135F and W105F and a double Trp mutant W135F/W105F have been characterized by steady state and time-resolved fluorescence and phosphorescence at 298 K and 77 K. Results indicate that Trp135 is more buried with less polar and more hydrophobic environment whereas the Trp105 is solvent exposed. Quantum yields (capital EF, Cyrillic) suggest that the singlet-singlet (S <--> S) non-radiative energy transfer (ET) from the Trp135 to the Trp105 occurs with 66% efficiency. The simulation of the fluorescence spectra of the WT and the time resolved studies support the results. Lifetime and capital EF, Cyrillic of the single Trp mutants suggest an intrinsic static quenching of the Trp105. The results at 77 K indicate that the ET takes place from the lowest triplet state (T(1)) of the Trp105 to the T(1) of the Trp135 apart from the backward S <--> S ET from the Trp105 to the Trp135. The triplet-triplet (T <--> T) ET implies a distance of <10 A between the Trp105 and the Trp135. Using the crystal structure of Vesicular Stomatitis Virus (VSV) phosphoprotein exhibiting about 34% similarity with the CHPV P protein, a homology modelling of CHPV supports the observed distance between the Trp residues, the S <--> S ET efficiency and the environments of the Trp residues in CHPV.

An Investigation of the Molecular Interactions of Diacetylcurcumin with Ribonuclease A

Curcumin is a natural product with diverse pharmacological activities. Studies of curcumin and its structural derivatives have been a subject of growing interest as a result of their diverse biological activities. We report the interaction of diacetylcurcumin (DAC) with Ribonuclease A (RNase A). The binding constant of DAC with RNase A was found to be of the order of 10(4) M(-1). The intrinsic fluorescence of RNase A was quenched by DAC with a quenching constant of 2.2 x10(4) M(-1). The distance between the fluorophore of RNase A and DAC was found to be 2.6 nm, calculated from a Förster type fluorescence resonance energy transfer (FRET). Secondary structural changes of RNase A after binding were analyzed from circular dichroism and Fourier transform infrared studies. Protein-ligand docking studies were conducted to determine the residues involved in the interaction of RNase A with DAC and changes in the accessible surface of the interacting residues were calculated accordingly.

Inherent Aggregation Propensity of Flanking Residues Attached to Polyglutamines: Implication to Aggregation Inhibition

Expansion of polyglutamine (polyQ) sequence in some proteins leads to their aggregation, which is responsible for neurodegenerative diseases like Huntingtons disease, ataxia etc. A flanking domain is usually fused at the N-terminal to polyQ in these proteins. On linking the flanking residues to polyQ, they accelerate aggregation of the proteins, which initiates from the flanking residues. In this report the inherent propensity of the flanking residues towards aggregation in six aggregating proteins has been elucidated from their primary sequences with the help of Betascan and PASTA programs and explored unambiguously. This will provide a molecular mechanism of this process. Suppression of aggregation using chaperones like αB-crystallin by masking the exposed hydrophobic surface of flanking residues is also documented through molecular docking, which could be applied for inhibition of aggregation of this type of proteins.