top of page


Our collaborative work on the "mPTX" molecule enhances sperm competence for IVF has been covered by the media.

IITH You tube:

Print or online media: Link to doc file ‘mPTX-IVF- News-Coverage’

Fig. 2: Schematic representation of designed molecule enhances the sperm competence (sperm motility, capacitation etc) for in vitro fertilization (IVF) through inhibiting Phosphodiesterases (PDE) which break cAMP to AMP.

Biochemistry journal cover page: Helical and β-turn conformations in the peptide recognition regions of the VIM1 PHD finger abrogate H3K4 peptide recognition



Characterization of allosteric regulation of PARP1 and PARP2 enzymes

Poly (ADP-ribose) polymerase 1 (PARP1) is an abundant nuclear enzyme, often referred to as “genome guardian”, along with PARP2, as it plays a very crucial role in maintaining the genome integrity. Enzyme activity of human PARP-1 and PARP2 are allosterically upregulated through binding to damaged DNA. There are Poly (ADP-ribose) polymerases (PARPs) inhibitors: FDA approved (Olaparib, Rucaparib and Niraparib) and under clinical trials (Talazoparib, Veliparib, CEP 9722 etc.), recommended to treat ovarian cancer, primary peritoneal cancer, breast cancer, prostate cancer, non-small-cell lung cancer, colorectal cancer etc. Major shortcoming of these PARP inhibitors for pharmacological use is, they are competitive inhibitors i.e., bind to the orthosteric site or the co-factor NAD+ binding site i.e., consequently can non-specifically bind to the active PARPs and also to many NAD+ and NADP+ dependent enzymes. Therefore, designing specific allosteric inhibitors against PARPs especially PARP1 and PARP2 would circumvent off target effect of above PARP inhibitors.

We unraveled the mechanism of allosteric regulation of DNA binding and catalytic activities of PARP1 by its catalytic product poly ADP ribose (PAR) polymer. Our work discovers dedicated PAR and DNA reader domains of the PARP1, and uncovers a novel mechanism of allosteric stimulation, but retardation of DNA-dependent activities of PARP1 by PAR (BioRXiv, 2021). In nutshell, PAR acts as both partial allosteric stimulator and partial allosteric inhibitor (partial antagonist) of PARP1 (Fig. 1), These studies would help in designing allosteric inhibitors/stimulators against PARP1 and PARP2.

image parp.png

Fig. 1. Schematic representation of proposed allosteric regulation of PARP1 by PAR and DNA. (A) Binding of PAR to ZnF3-WGR domains may lead to spatial reorganization of PARP1 domains to get into certain conformation that stimulates the catalytic activity of PARP1. (B) DNA binding induces global conformational changes in the protein that leads to its maximal activation. (C) Binding of PAR and DNA to PARP1 might be non-competitive reversible that may lead to two different conformational outcomes, resulting in differential catalytic stimulation, which could be an average of PAR- and DNA-dependent catalytic stimulation of the PARP1. The displacement of DNA by the PAR from the DNA-PARP1 complex eventually retards the DNA dependent stimulation but at the same time PAR binding itself stimulate PARP1 activity, therefore the activation is modulated by cross talk between DNA and PAR binding to PARP1.

Drug Design
Engineering phosphodiesterase inhibitors to modulate sperm functions for In vitro fertilization (IVF)

Phosphodiesterases are the enzymes that hydrolyze the cyclic nucleotides and thus regulate their cellular levels. We have redesigned and biochemically evaluated phosphodiesterase inhibitor drug pentoxifylline (PTX) analogs against phosphodiesterases functioning in sperms. PTX is one of the widely used pharmacological agents for aiding viable sperm selection in asthenozoospermic patients and to increase sperm motility prior to intracytoplasmic sperm injection, but it induces premature acrosome reaction. Ex vivo evaluation of one of the designed PTX analogs on sperm by our collaborator (Prof. Guruprasad Kalthur, Kasturba Medical College, Manipal Academy of Higher Education) has shown improved human sperm motility and longevity and reduced pre-acrosomal reaction compared to the parent molecule (Nature Scientific Reports, 2021)  {Fig. 2 }.

Vaccine design

Designing ferritin nanocage based vaccine candidates for SARS-CoV-2

Using the homotrimeric ‘6-helical bundle’ of the spike protein and other peptide epitopes of SARS-CoV-2, we designed a chimeric ferritin nanocage. Designed multivalent and multiple epitopes' protein nanocage and scaffold could mount both humoral and T-cell mediated immune response against SARS-CoV-2 (JBSD, 2022).

Characterization of Host Receptor Interaction with Envelop Protein of Kyasanur Forest Disease Virus and Predicting Suitable Epitopes for Vaccine Candidate (Under Review)

  • Editor-in-Chief choice to feature on the electronic front cover of the issue of The FEBS Journal

  • Editor's Choice paper for volume 290, issue 21 of The FEBS Journal.

  • This article is considered to promote the new issue on our Twitter feed and LinkedIn 

  • Considered for free open access: to maximise its readership

febs 2.jpg

 Schematic representation of designing of SARS-CoV-2’s epitope peptides mounted ferritin (FR) nanocage as vaccine candidate.

Epigenetic Regulation

We aim to understand how chemical tags/barcodes on histones and DNA are established (written), removed (erased) and/ or recognized (read) by the enzymatic machineries and/ or modular proteins.

Mechanistic insights into allosteric regulation of methylated DNA and histone H3 recognition by SRA and SET domains of SUVH5 and the basis for di-methylation of lysine residue

Su-(var)3-9 homologue 5 (SUVH5), a member of SUVH family of histone lysine methyltransferase (HKMT) in Arabidopsis is involved in epigenetic regulation of chromatin by recognizing 5-methyl-cytosine (5mC), in both CpG and non-CpG DNA context, through SRA domain and simultaneously performing the di-methylation of lysine 9 of histone 3 (H3K9) through SET domain. Here, we establish that the SET domain of SUVH5 allosterically restricts the SRA domain to the 5mC containing strand(s) of fully-methylated CpG, hemi-methylated CpG and methylated CpHpH DNA (Under review) (Fig. 4). In addition, SET domain enhances the binding affinity of the SRA-SET dual domains to fully-mCpG but not to hemi-mCpG. Further, computational analyses and QM/MM calculations explain the bases for robust mono-MTase but weak di-MTase activities of SUVH5 (FEBS J. 2022). Given that the majority of eukaryotic proteins, including those involved in epigenetic gene regulation, contain more than one domain, our study suggests that understanding the allosteric regulation among multiple domains of proteins is relevant for unraveling biological outcomes.

Mechanistic insights into combinatorial recognition of H3R2 and H3K9me2 barcodes by Dual Domains (TTD-PHD) of UHRF1

Biochemical (binding) and dynamics (MD simulation) studies elucidate that the TTD (Tandem Tudor Domain) of UHRF1 binds to dimethylated lysine 9 of histone H3 and subtle variations of key residues at the binding pocket determine the status specific recognition of methylated lysines (Fig. 5) by the histone mark reader domains of UHRF1 (Biochimie, 2018).

Fig. 4. Schematic representation of regulation of SRA and SET domains binding to methylated DNA and H3 peptide. Top panel: Representation showing that the SET domain of SUVH5 allosterically regulates the recognition of different methylated DNA by SRA domain. The red dots represent 5mC base. The stoichiometry of protein to DNA duplex is mentioned above the arrows. Bottom panel: Representation showing that recognition of fully-mCpG DNA by SRA allosterically regulates the binding of H3 peptide to the SET domain of SUVH5. The representation on the bottom left shows that SET domain recognizes the conserved “A(R/K)KST” motif in the histones H3 and H2A variants-HTA2 and HTA13. The positively substitutes residue in the motif is highlighted in yellow and the target lysine is shown in red font.


Fig. 5: Isothermal Titration Calorimetry (ITC) binding studies show the combinatorial binding of epigenetic (H3R2 and H3K9me) marks, and preferential recognition of H3K9me2 mark over H3K9me3 mark by the TTD-PHD dual domains of UHRF1.

UHRF1-SRA recognizes symmetric non-CG methylated DNA through dual-flip out of 5-methyl cytosines

UHRF1, is a bonafide reader of hemi-methylated DNA and essential for the maintenance of DNA methylation. Using ITC binding and X-ray crystallographic structural studies we have shown that SRA domain of UHRF1 can recognize different methylation status of DNA and single base spacer between symmetric 5mCs is required for dual flip out recognition of 5mCs in non-CG context (Fig. 6) (Int J Biol Macromol, 2021).


Fig. 6: X-ray crystallography structure of UHRF1 SRA recognizes cytosine-methylated DNA in CHG sequence context.

HMGB proteins from protozoa pathogens

High mobility group (HMG) proteins, are non-histone chromatin architectural proteins, bind different DNA structures and chromatin, induce conformational changes in the chromatin and topological changes in DNA that facilitate the replication, transcription, recombination and repair of both nuclear and mitochondrial DNA.

Non-canonical DNA binding specificities of KAP6 from Trypanosoma causal agent of sleeping sickness

Our investigations revealed that KAP6 binds non-canonical DNAs (splayed and flap DNA, Holliday Junction) tighter than B-form DNA. Simulation analyses revealed that the ~90° bend in DNA induced by KAP6 HMG box is a result of two ~45° bends, by helices of the protein (Fig. 7). Our data also suggests that the orthologs of KAP6 are oligomers in solution, which could be necessary for their functioning such as 180° DNA bending and looping during kDNA packaging (Int J Biol Macromol, 2020).


Fig. 7: Biochemical and computational (MD simulation and metadynamics) analyses suggest that HMGB box of TtKAP6 protein bent the DNA by 90°, which is required for packing of mitochondrial DNA of Trypanosoma.

Characterization of substrate and product specificities of histone lysine Methyltransferases from arabidopsis, rice and maize

Methylation of lysine by histone lysine methyltransferases (HKMTs) has been implicated in regulation of gene expression. To unravel histone substrate specificity, degree of methylation and catalytic activity, we analyzed Arabidopsis Trithorax-like protein (ATX), Su(var)3-9 homologs protein (SUVH), Su(var)3-9 related protein (SUVR), ATXR5, ATXR6, and E(Z) HKMTs from Arabidopsis, maize and rice through sequence and structure comparison. We proposed that subtle variations of key residues at substrate or SAM (S-Adenosylmethionine) binding pocket, around the catalytic pocket, or presence of pre-SET and post-SET domains in HKMTs of the aforementioned plant species lead to variations in class-specific HKMT functions and further determine their substrate specificity, the degree of methylation and catalytic activity (Proteins, 2018).


Mechanistic insights into combinatorial recognition of H3R2 and H3K9me2 barcodes by Dual Domains (TTD-PHD) of UHRF1

Biochemical (binding) and dynamics (MD simulation) studies elucidate that the TTD (Tandem Tudor Domain) of UHRF1 binds to dimethylated lysine 9 of histone H3 and subtle variations of key residues at the binding pocket determine the status specific recognition of methylated lysines (Figure 3) by the histone mark reader domains of UHRF1 (Biochimie, 2018).

Mechanistic insights into the recognition of 5-methylcytosine oxidation derivatives by the SUVH5 SRA domain

Our Isothermal Titration Calorimetry (ITC) binding and X-ray crystallographic structural studies unraveled the molecular basis of the recognition of the 5mC oxidation derivatives ( 5-hydroxymethyl (5hm), 5-carboxymethyl (5ca) and 5-formylmethyl (5fc) in the CG sequence context by the SET- and RING-associated domain (SRA) of the SUVH5 (SUVH5 SRA).We reported the 2.6 Å resolution crystal structure of the SUVH5 SRA domain in a complex with fully hydroxymethyl-CG (5hmCG) and demonstrated a dual flip-out mechanism, whereby the symmetrical 5hmCs are simultaneously extruded from the partner strands of the DNA duplex and are positioned within the binding pockets of individual SRA domains. This study demonstrated that the SRA domain binds to the 5hmCG DNA duplex in a manner similar to methylated CG (5mCG) (Nature Scientific Reports, 2016).

Enzyme Engineering
Enzyme characterization or engineering for active pharmaceutical ingredients synthesis

Synthesis of single enantiomer of active pharmaceutical ingredients (APIs) or drugs has always been a challenge and of great demand in pharmaceutical industry, since safety and efficacy of many drug products are dependent on their chirality. Even active ingredients of significant number of chemicals used as pesticides, herbicides or plant growth regulators in agricultural industry are single enantiomers. Owing to high enantioselectivity and ‘greener process’, enzymes are gaining increasing attention as catalysts in the chemical and pharmaceutical industry for asymmetric synthesis (ACS Chemical Biology, 2022). Aim of this project is to characterize or engineer epoxide hydrolases, imine reductases and transaminases of fungal, bacterial or plant origin for asymmetric synthesis of enantiopure compounds.


Mechanistic understanding and engineering of Strictosidine synthase for improved enzyme properties

3-α-(S)-strictosidine is the substrate for various metabolic pathways in plants that synthesize nearly 2000 monoterpene indole alkaloids including pharmaceutical compounds (camptothecin, ajmaline, vindoline, quinine, etc) used to treat cancer, malaria, hypertension or schizophrenia etc. 3-α-(S)-strictosidine is a complex stereoisomer and its chemical synthesis is challenging. Therefore, we want to engineer the enzyme Strictosidine synthase for broad substrate specificity to synthesize variants of strictosidine, and to enhance its catalytic efficiency to increase the productivity and yield of 3-α-(S)-strictosidine and its variants (In collaboration with Prof. Dr. Ulrich Schwaneberg, Chair for Biotechnology, RWTH Aachen University, Germany).

bottom of page