Research Associate (Apr 2014 - Mar 2015)

Advisor: Prof. Dipak Dasgupta

● Investigated the kinetics preference of small molecule binding to quadruplex over duplex DNA if any

● Investigated the interaction of small molecules with histones and study their effect on histone modification marks

Doctoral Work (Aug 2009 - Mar 2014)

Title: Effect of the Plant Alkaloids on Structure and Function of G-quadruplex DNA

Advisor: Prof. Dipak Dasgupta

Work Synopsis: During my Ph.D. tenure I have focused on the interaction of plant alkaloids with therapeutic potential with G-quadruplex DNA, which is found in the telomeric region of chromosomes and in the transcription regulatory region of various oncogenes. Using various biophysical and biochemical techniques.

I have characterized and quantitated the interaction of two G-quadruplex forming sequences with three plant alkaloids namely- ellipticine (ELP), sanguinarine (SGR) and chelerythrine (CHL). The two quadruplex sequences used for the studies are – the human telomeric DNA sequence (H24) and the NHEIII upstream of the promoter of the c-myc oncogene (Pu27). Under the experimental conditions, the H24 adopts a (3+1) mixed hybrid structure and Pu27 adopts an all parallel structure.

In addition to the biophysical techniques used to characterize the interaction of the small molecules with quadruplex DNA in vitro, TRAP assay was carried out with cell extracts from breast cancer cell lines MDAMB-231 to study the effect of these molecules on telomerase activity. Furthermore the ability of SGR and CHL to block hybridization of Pu27 with its complementary strand in a concentration dependent manner has also been studied. Control experiment with a sequence which does not fold into a quadruplex structure confirms that the inhibition is due to stabilization of the quadruplex structure by SGR and CHL.

The affinity of all three molecules towards quadruplex DNA, corresponding duplex DNA and a random DNA sequence, ct DNA has also been compared.

The above studies led us to conclude that small planar molecules can effectively interact with quadruplex DNA in vitro and stabilize them. These molecules can also effectively block the activity of telomerase and Pu27 hybridization in a concentration dependent manner. These molecules, therefore, has the potential to act as putative anticancer agents. Using these small molecules as the starting point, other molecules with greater specificity for quadruplex structures and lower cellular toxicity can also be synthesized which can emerge as potential anticancer agents.

Related Publications:

  1. Ghosh S, Dasgupta D. (2015) Quadruplex forming promoter region of c-myc oncogene as a potential target for a telomerase inhibitory plant alkaloid, chelerythrine. Biochem Biophys Res Commun 459 (1) 75-80
  2. Ghosh S, Jana J, Kar R, Chatterjee S, Dasgupta D. (2015) Plant alkaloid chelerythrine induced aggregation of human telomere sequence-an unique mode of association between a small molecule and a quadruplex. Biochemistry 54 (4) 974-986
  3. Ghosh S, Kar A, Chowdhury S, Dasgupta D. (2013) Ellipticine binds to human telomere sequence – an additional mode of action as a putative anticancer agent? Biochemistry 52 (24) 4127-4137
  4. Ghosh S, Pradhan SK, Kar A, Chowdhury S, Dasgupta D. (2013) Molecular basis of recognition of quadruplexes human telomere and c-myc promoter by the putative anticancer agent sanguinarine. Biochim Biophys Acta 1830 (8) 4189-4201

Other Research Work during Doctoral Period (Aug 2009 - Mar 2014)

Advisor: Prof. Dipak Dasgupta

  • Investigated the effect of groove width on the binding of an intercalator and groove binder with duplex DNA.
  • Investigated the interaction of bivalent copper ion with the anticancer antibiotic Chromomycin A3 using spectroscopic techniques (including NMR data analysis).
  • Investigated the thermal stability of lamin Ig fold domain and its mutant.
  • Investigated the thermal stability of chimeric proteins.

Related Publications:

  1. Bera M, Kothamarthi HC, Dutta S, Ray A, Ghosh S, Bhattacharyya D, Ainavarapu SR, Sengupta K. (2014) Characterization of unfolding mechanism of human lamin A Ig fold by single-molecule force spectroscopy – Implications in EDMD. Biochemistry 53 (46) 7247-7258
  2. Ray S, Blaise M, Roy B, Ghosh S, Kern D, Banerjee R. (2014) Fusion with Anticodon Binding Domain of GluRS is Not Sufficient to Alter the Substrate Specificity of a Chimeric Glu-Q-RS. Protein J 33 (1) 48-60
  3. Das S, Ghosh S, Dasgupta D, Sen U, Mukhopadhyay D. (2012) Biophysical studies with AICD-47 reveal unique binding behavior characteristic of an unfolded domain. Biochem Biophys Res Commun 425 (2) 201-206
  4. Lahiri S, Takao T, Devi PG, Ghosh S, Ghosh A, Dasgupta A, Dasgupta D. (2012) Associatio of aureolic acid antibiotic, chromomycin A3 with Cu+2 and its negative effect upon DNA binding property of the antibiotic. Biometals 25 (2) 435-450

Pre-doctoral Work (Post M.Sc. Project: Apr 2009 - June 2009)

Title: Unusual DNA Structures as Targets for Small Compounds with Therapeutic Potential

Advisor: Prof. Dipak Dasgupta

Work Synopsis: It is evident that the original B-duplex DNA discovered by Watson and Crick is not the only structure that DNA can adopt. Along with A- and Z- forms of duplex DNA, DNA can also form triplexes, G-quadruplexes (formed in G-rich regions), i-motifs (formed in C-rich regions) and PNAs which fall under the unusual DNA structures. Of all these unusual forms that DNA can adopt, in the recent years G-quadruplex has attracted the attention of researchers the most on account of their biological importance. G-quadruplexes are found in telomeric regions, promoter and immunoglobulin switch regions and recombination hot spots. They represent a potential drug

target for DNA-binding compounds. Researchers over the past few years have targeted G-quadruplex DNA with small molecules to evaluate their therapeutic potentials. In this review, I had focused on

  • The structural diversity of G-quadruplex DNA
  • The role of G-quadruplex DNA in vivo
  • Small molecules that targets and stabilizes G-quadruplexes, and in turn possess potential therapeutic value and
  • The role of quadruplex aptamers as potential therapeutic agents.

This review also provides an overview of the various techniques that are available to study the structure of quadruplex DNA and their interaction with small molecules with therapeutic potential.

Related Publication:

  1. Ghosh S, Majumder P, Pradhan SK, Dasgupta D. (2010) Mechanism of interaction of small transcription inhibitors with DNA in the context of chromatin & telomere. BBA - Gene Regulatory Mechanisms 1799 (10-12) 795-809

Pre-doctoral Work (M.Sc. Project: Aug 2007 - Mar 2008)

Title: Investigation of the Interaction between 1-anthracene sulphonate and Transport Proteins

Advisor: Prof. Subhash Chandra Bhattacharya, Chemistry Department, Jadavpur University, Kolkata, INDIA

Work Synopsis: The interaction of 1-anthracene sulphonate (1-AS) with model proteins, bovine serum albumin (BSA) and human serum albumin (HSA) containing 2 and 1 tryptophan residues respectively, was studied employing steady state and time resolved fluorescence techniques. The aim of the study was to utilize the fluorescence properties of 1-AS and understand its interaction with biological targets, in this case BSA and HSA. The results indicated a marked change in fluorescence property of BSA and HSA upon binding with 1-AS.

The fluorescence spectrum of BSA and HSA was quenched on interaction with 1-AS with the appearance of an isoemissive point. Linearity of Stern-Volmer plot for both proteins indicated that a single type of quenching occurs in both systems. Since the absorption spectrum of BSA/HSA did not change on addition of 1-AS, it was concluded that static quenching doesnot occur in the system. Analysis of the binding data revealed that binding stoichiometry is 1:1 and BSA has a higher affinity for 1-AS than HSA.

Fluorescence Resonance Energy Transfer (FRET) between excited tryptophan in the transport proteins and 1-AS, was used for studying the relaxation dynamics of biological molecules. Results suggested that the efficiency of energy transfer was higher in BSA than in HSA.

Time resolved fluorescence spectroscopy (TCSPC) was done to obtain an idea about the change in the microenvironment of 1-AS upon binding to the proteins. The average fluorescence lifetime of BSA/HSA decreased with increase in 1-AS concentration. This observation correlated with

the steady state fluorescence data that the quenching of BSA/HSA by 1-AS was due to FRET by binding of 1-AS to the tryptophan moieties present in the protein.

It can be concluded from the above studies that, FRET based techniques could be used to study the interaction of fluorescent probes with proteins. It could also find application in labeling damaged proteins through their interaction with bioactive compounds.

Related Publication:

  1. Banerjee P, Ghosh S, Sarkar A, Bhattacharya SC. (2011) Fluorescence resonance energy transfer: A promising tool for investigation of the interaction between 1-anthracene sulphonate and serum albumins. J. Lumin. 131 (2) 316-321