Qualification: 
Ph.D, MSc, M.Tech
nanditamishra@am.amrita.edu

Dr. Nandita Mishra currently serves as Associate Professor at the School of Biotechnology, Amrita Vishwa Vidyapeetham. Ph. D. in Biotechnology, IIT Kharagpur in 2005. Dr. Mishra joins us from University of Texas Health Science Centre, San Antonio, USA.

Education

  • Ph.D. Biotechnology, Indian Institute of Technology, Kharagpur.
  • M.Tech. Biotechnology and Engineering, Indian Institute of Technology, Kharagpur.
  • M.Sc. Life Sciences, Sambalpur University, Orissa.
  • B.Sc. Botany (Honors) Sambalpur University, Orissa.
     

Research Interests

Awards and Scholarships

  • Awarded Fellowship from Govt. of India for Doctoral studies.
  • Ministry of Human Resources & Dev, Government of India fellowship for M. Tech. studies.
  • Qualified Graduate Aptitude Test in Engineering (GATE).
  • Visiting Research fellow, University of Guelph, Canada (2005) with Prof. Manish Raizada.
  • Postdoctoral fellowship, University of Texas Health Science Center at San Antonio, Texas, USA with Prof. Pothana Saikumar (2006-2010).
  • Awarded Scholarship from Government of Orissa for Excellence in Secondary Examination.
  • Award of Excellence from Amrita Vishwa Vidyapeetham (2012-13)

Teaching

  1. BIO317/Research Methodology/B.Sc. Biotechnology and B.Sc. Microbiology/2 Credits/Semester 5
  2. BIO399/ Project /B.Sc. Biotechnology and B.Sc. Microbiology/10 Credits/Semester 6
  3. BIO401/Molecular Biology/M.Sc. Biotechnology and M.Sc. Microbiology/3 Credits/Semester 1
  4. BIO497/Research Methodology/M.Sc. Biotechnology and M.Sc. Microbiology/2 Credits/Semester 1
  5. BIO408/Recombinant DNA Technology/M.Sc. Biotechnology and M.Sc. Microbiology/3  Credits/Semester 2
  6. BIO481/Recombinant DNA Technology Lab/M.Sc. Biotechnology and M.Sc. Microbiology/2  Credits/Semester 2
  7. BIO517/ Plant Biotechnology/ M.Sc. Biotechnology /3 Credits/ Semester 2
  8. BIO588/Cell & Molecular Biology Lab/M.Sc. Biotechnology & M.Sc. Microbiology/2  Credits/ Semester 3/
  9. BIO599/Project/M.Sc. Biotechnology & M.Sc. Microbiology/10 Credits/Semester 4
  10. BIO464/ Cancer Biology Elective/ MSc Biotechnology/MSc Microbiology/4 Credits/Semester 3
     

Grants received

Publications

Publication Type: Journal Article

Year of Publication Title

2020

S. Khatun, Biswas, S., Anupama Binoy, Podder, A., Dr. Nandita Mishra, and Bhuniya, S., “Highly chemoselective turn-on fluorescent probe for ferrous (Fe2+) ion detection in cosmetics and live cells”, Journal of Photochemistry and Photobiology B: Biology, vol. 209, p. 111943, 2020.[Abstract]


In modern society, the use of cosmetics has increased extensively; unfortunately, so-called several toxic metal salts are present as the colorant or filler in cosmetics. The ferrous ion (Fe2+) is one of the metal ions used in cosmetics as a colorant. Ferrous ion (Fe2+) is a vital component in live cells. Considering the adverse effect of high doses of ferrous ions in cosmetics and live cells, we developed a turn-on fluorescent probe PFe(II) for quantitative estimation of ferrous ion (Fe2+) in cosmetics and monitoring of labile ferrous (Fe2+) ion in live cells. The fluorescent probe PFe(II) showed a visual color change from colorless to orange in the presence of ferrous ion (Fe2+) in the cosmetics. We observed that UV-absorption increased at 390 nm upon incubation with ferrous ion (Fe2+). The probe PFe(II) has provided quantitative information on ferrous ion (Fe2+) in various cosmetics, kajol, lip balm, face foundation, mascara, eyeliner, lipliner, face makeup, sindoor, lipstick, nail polish in ppm level through the fluorescence signaling at 460 nm.The probe PFe(II) provided information on labile Fe2+ ion pool via a fluorescence imaging. It is a new addition to the diagnostic inventory for detecting ferrous ion in live cells and cosmetics.

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2019

Damu Sunilkumar, Drishya G., Chandrasekharan, A., Shaji, S. K., Chinchu Bose, Jossart, J., J. Jefferson P. Perry, Dr. Nandita Mishra, Dr. Geetha Kumar, and Dr. Bipin G. Nair, “Oxyresveratrol drives caspase-independent apoptosis-like cell death in MDA-MB-231 breast cancer cells through the induction of ROS.”, Biochem Pharmacol, p. 113724, 2019.[Abstract]


Earlier studies from our laboratory have demonstrated that Oxyresveratrol (OXY), a hydroxyl-substituted stilbene, exhibits potent inhibition of human melanoma cell proliferation. The present study defines a cytotoxic effect of OXY on the highly chemo-resistant, triple-negative human breast cancer cell line MDA-MB-231. OXY-mediated cell death resulted in accumulation of cells at the sub-G1 phase of the cell cycle, induced chromatin condensation, DNA fragmentation, phosphatidylserine externalization and PARP cleavage, indicative of apoptosis. Interestingly, morphology and cell viability studies with the pan-caspase inhibitor, QVD-OPH revealed that OXY-induced cell death was caspase-independent.Docking studies also showed that OXY can bind to the S1 site of caspase-3, and could also exert an inhibitory effect on this executioner caspase. The immunoblot analysis demonstrating the absence of caspase cleavage during cell death further confirmed these findings. OXY was also observed to induce the production of reactive oxygen species, which caused the depolarization of the mitochondrial membrane resulting in translocation of Apoptosis Inducing Factor (AIF) into the nucleus. Pretreatment of the cells with N-Acetyl Cysteine antioxidant prevented cell death resulting from OXY treatment. Thus, OXY initiates ROS-mediated, apoptosis-like cell death, involving mitochondrial membrane depolarization, translocation of AIF into the nucleus, and DNA fragmentation, resulting in caspase-independent cell death in MDA-MB-231 cells. The cytotoxicity manifested by OXY was also observed in 3D cell culture models and primary cells, thereby providing a basis for the utilization of OXY as a novel template for the future design of anticancer therapeutics.

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2019

Divya Nedungadi, Anupama Binoy, Vinod, V., Muralidharan Vanuopadath, Sudarslal Sadasivan Nair, Dr. Bipin G. Nair, and Dr. Nandita Mishra, “Ginger extract activates caspase independent paraptosis in cancer cells via ER stress, mitochondrial dysfunction, AIF translocation and DNA damage”, Nutrition and Cancer, pp. 1-13, 2019.[Abstract]


AbstractThe rhizome of ginger (Zingiber officinale) a common culinary agent is also known for its medicinal activity. We have earlier reported that pure 6-shogaol, an important component of ginger induces paraptosis in triple negative breast cancer (MDA-MB-231) and non small cell lung (A549) cancer cells. However, the chemopreventive potential of the whole ginger extract in food remains to be elucidated. Here, we demonstrate for the first time that ginger extract (GE) triggers similar anticancer activity/paraptosis against the same cell lines but through different molecular mechanisms. Q-TOF LC-MS analysis of the extract showed the presence of several other metabolites along with 6-shogaol and 6-gingerol. GE induces cytoplasmic vacuolation through ER stress and dilation of the ER. Drastic decrease in the mitochondrial membrane potential and ATP production along with the excess generation of ROS contributed to mitochondrial dysfunction. Consequently, GE caused the translocation of apoptosis inducing factor to the nucleus leading to the fragmentation of DNA. Taken together, these show a novel mechanism for ginger extract induced cancer cell death that can be of potential interest for cancer preventive strategies.

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2019

Anupama Binoy, Divya Nedungadi, Katiyar, N., Chinchu Bose, Dr. Sahadev Shankarappa, Dr. Bipin G. Nair, and Dr. Nandita Mishra, “Plumbagin induces paraptosis in cancer cells by disrupting the sulfhydryl homeostasis and proteasomal function”, Chemico-Biological Interactions, p. 108733, 2019.[Abstract]


Plumbagin (PLB) is an active secondary metabolite extracted from the roots of Plumbago rosea. In this study, we report that plumbagin effectively induces paraptosis by triggering extensive cytoplasmic vacuolation followed by cell death in triple negative breast cancer cells (MDA-MB-231), cervical cancer cells (HeLa) and non-small lung cancer cells (A549) but not in normal lung fibroblast cells (WI-38). The vacuoles originated from the dilation of the endoplasmic reticulum (ER) and were found to be empty. The cell death induced by plumbagin was neither apoptotic nor autophagic. Plumbagin induced ER stress mainly by inhibiting the chymotrypsin-like activity of 26S proteasome as also evident from the accumulation of polyubiquitinated proteins. The vacuolation and cell death were found to be independent of reactive oxygen species generation but was effectively inhibited by thiol antioxidant suggesting that plumbagin could modify the sulfur homeostasis in the cellular milieu. Plumbagin also resulted in a decrease in mitochondrial membrane potential eventually decreasing the ATP production. This is the first study to show that Plumbagin induces paraptosis through proteasome inhibition and disruption of sulfhydryl homeostasis and thus further opens up the lead molecule to potential therapeutic strategies for apoptosis-resistant cancers.

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2019

K. Naidu Bobba, Anupama Binoy, Koo, S., Nedungadi, D., Podder, A., Sharma, A., Dr. Nandita Mishra, Kim, J. Seung, and Bhuniya, S., “Direct readout protonophore induced selective uncoupling and dysfunction of individual mitochondria within cancer cells”, Chemical Communications, vol. 55, no. 45, pp. 6429-6432, 2019.[Abstract]


Concurrently, manipulation of mitochondrial activity and its monitoring have enormous significance in cancer therapy and diagnosis. In this context, a fluorescent probe MitoDP has been developed for validating H2S mediated protonophore (2,4-dinitrophenol, DNP) induced mitochondrial membrane potential change, ROS formation and ATP depletion in cancer cells. The extent of protonophore activation for mitochondrial dysfunction is monitored through fluorescence signalling at 450 nm. The current study provides a proof for the concept of endogenous H2S-mediated controlled and spatial release of bioactive agents, or toxins specifically in mitochondria of cancer cells.

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2018

P. Sagitha, Reshmi, C. R., Sundaran, S. P., Anupama Binoy, Dr. Nandita Mishra, and Sujith, A., “In-vitro evaluation on drug release kinetics and antibacterial activity of dextran modified polyurethane fibrous membrane.”, International Journal of Biological Macromolecules, vol. 126, pp. 717-730, 2018.[Abstract]


pH stimuli drug release nanofibrous membranes of polyurethane/dextran were developed for tailoring of antibacterial wound dressings. Incorporation of dextran in polyurethane (PU) showed increment in hydrophilicity, vapour transmission rate, percentage sorption values, and biodegradability. Dextran also acts as reinforcement filler in PU matrix. Dextran induces a high degree of platelet adhesion and hemostasis potential which is essential for promoting the wound healing process. Moreover, 20 wt% dextran loaded membranes (PU/20D) exhibited enhanced cell proliferation, attachment and viability against 3T3 fibroblasts. Curcumin loaded PU/20 dextran membrane exhibited pH-controlled drug release potency and synergistic antibacterial activity against gram-positive bacteria. It is confirmed that, PU/20D membranes could promote, pH-controlled drug release and synergistic antibacterial activity for a promising wound dressing material.

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2018

R. Rashmi, Divya Nedungadi, Podder, A., Dr. Nandita Mishra, and Bhuniya, S., “Monitoring of topoisomerase (I) inhibitor camptothecin release from endogenous redox-stimulated GO-polymer hybrid carrier”, Journal of Photochemistry and Photobiology B: Biology, vol. 189, pp. 14-20, 2018.[Abstract]


We have developed endogenous redox-responsive polymer conjugated GO-based hybrid nanomaterials (GO-PEGssFol-CPT) for delivery of anticancer drug camptothecin (CPT) to the cancer cells. The synthesized intermediate (PEGFol) and CPT loaded GO- PEGFol were characterized using Fourier transform infrared spectroscopy (FTIR) and H NMR. The morphological feature changes of TEM and AFM images have confirmed the loading of CPT on the nanocarrier and its release from the nanocarrier. The amount of CPT was loaded was found to be 14.2%. The extent of camptothecin (CPT) release from GO-BiotinPVA-CPT in the presence of different concentrations of glutathione (GSH) was monitored with the increase in the fluorescence intensity at λ 438 nm and UV-Vis absorbance at 366 nm. The time-dependent camptothecin (CPT) release was monitored in the presence of GSH. It was noticed that CPT was completely released from GO-PEGssFol-CPT within 45 min. This release process is free from interference by other ubiquitous analytes in the living system. The constant fluorescence intensity of GO-PEGssFol-CPT against acidic pH indicated that CPT would not be released in the extracellular region of cancer cells. Therefore, such delivery system could be used to prevent unwanted cytotoxicity to the healthy cells. The GO-PEGssFol-CPT showed higher antiproliferative activity against cervical cancer cells compared to the CPT. Thus, GO-PEGssFol-CPT can be a new material to deliver the anticancer drug to the target tumor region.

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2018

C. Porayath, M. K. Suresh, Dr. Raja Biswas, Dr. Bipin G. Nair, Dr. Nandita Mishra, and Dr. Sanjay Pal, “Autolysin mediated adherence of Staphylococcus aureus with Fibronectin, Gelatin and Heparin.”, International Journal of Biological Macromolecules, vol. 110, pp. 179-184, 2018.[Abstract]


Major autolysin (Atl) of Staphylococcus aureusis a cell surface associated peptidoglycan hydrolase with amidase and glucosaminidase domains. Atl enzymes (amidase and glucosaminidase) are known to participate in biofilm formation and also can bind with host matrices. Earlier studies demonstrated the binding of Atlwithfibronectin, thrombospondin 1, vitronectin and heat shock cognate protein Hsc70. Here, we have shown, Atl mediates attachment of S.aureus to heparin and gelatine as well. The atl mutant strain demonstrated around 2.5 fold decreased adherence with fibronectin, gelatin and heparin coated microtiter plates. The microscopic studies confirmed the reduced binding of atl mutant with them compared to its parental wild type and complemented mutant strains. Amidase and glucosaminidase were expressed as N-terminal histidine tagged proteins from Escherichia coli, purified and refolded. We found refolded amidase bind with fibronectin, gelatin and heparin; whereas refolded glucosaminidase binds with only fibronectin and heparin but not gelatin. These results reemphasize Atl as one of the crucial proteins from Staphylococcus that facilitate their binding with multiple host cellular components during colonization and infection.

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2018

D. Nedungadi, Binoy, A., Pandurangan, N., Pal, S., Nair, B. G., and Dr. Nandita Mishra, “6-Shogaol induces caspase-independent paraptosis in cancer cells via proteasomal inhibition”, Experimental Cell Research, vol. 364, no. 2, pp. 243-251, 2018.[Abstract]


An α, β-unsaturated carbonyl compound of ginger, 6-Shogaol (6S), induced extensive cytoplasmic vacuolation and cell death in breast cancer cell (MDA-MB-231) and non-small lung cancer (A549) cells. In the presence of autophagic inhibitors the cells continued to exhibit cytoplasmic vacuolation and cell death clearly distinguishing it from the classic autophagic process. 6S induced death did not exhibit the characteristic apoptotic features like caspase cleavage, phosphatidyl serine exposure and DNA fragmentation. The immunofluorescence with the Endoplasmic Reticulum (ER) resident protein, calreticulin indicated that the vacuoles were of ER origin, typical of paraptosis. This was supported by the increase in level of microtubule associated protein light chain 3B (LC3 I and LC3 II) and polyubiquitin binding protein, p62. The level of ER stress markers like polyubiquitinated proteins, Bip and CHOP also consistently increased. We have found that 6S inhibits the 26S proteasome. The proteasomal inhibitory activity was elucidated by a) molecular docking of 6S onto the active site of β5 subunit and b) reduced fluorescence by the fluorogenic substrate of the chymotrypsin-like subunit. In conclusion these studies demonstrate for the first time that proteasomal inhibition by 6S induces cell death via paraptosis. So 6-shogaol may act as a template for anti-cancer lead discovery against the apoptosis resistant cancer cells.

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2017

N. Velusamy, Anupama Binoy, Bobba, K. Naidu, Divya Nedungadi, Dr. Nandita Mishra, and Bhuniya, S., “A bioorthogonal fluorescent probe for mitochondrial hydrogen sulfide: new strategy for cancer cell labeling”, Chem Commun (Camb)., vol. 53, no. 62, pp. 8802-8805, 2017.[Abstract]


We report the application of a chemodosimeter {'}turn on{'} fluorescent probe for detecting endogenous H2S formation in cancer cells. Mito-HS showed a bathochromic shift in the UV-vis-absorption spectrum from 355 nm to 395 nm in the presence of H2S. Furthermore{,} it showed an [similar]43-fold fluorescence enhancement at [small lambda]em = 450 nm in the presence of H2S (200 [small mu ]M). The cancer cell-specific fluorescence imaging reveals that Mito-HS has the ability to distinguish cancer cells from normal cells based on the level of endogenous H2S formation. In due course{,} Mito-HS would be a powerful cancer biomarker based on its ability to estimate endogenous H2S formation in living cells.

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2017

C. R. Reshmi, Menon, T., Anupama Binoy, Dr. Nandita Mishra, Elyas, K. K., and Sujith, A., “Poly(L-lactide-co-caprolactone)/collagen electrospun mat: Potential for wound dressing and controlled drug delivery”, International Journal of Polymeric Materials and Polymeric Biomaterials, vol. 66, no. 13, pp. 645-657, 2017.[Abstract]


Here we report a novel bioactive electrospun mat based on poly(L-lactide-co-caprolactone) (PLLC) and collagen for wound dressing and sustained drug delivery of gentamicin. PLLC/collagen electrospun mat loaded with 10% gentamicin showed bioactivity for 15 days against Gram-positive and Gram-negative bacteria. The in vitro cell culture of 3T3 fibroblasts confirmed that these electrospun mat provide an increased specific interface area and hydrophilicity to enhance cell attachment, proliferation, and migration. The modified PLLC/collagen mat provided an excellent enhancement in properties of antibacterial wound dressings with a minimum in vitro toxicity and high potency for promoting wound healing stages. © 2017 Taylor & Francis.

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2010

R. Kar, Dr. Nandita Mishra, Singha, P. K., Venkatachalam, M. A., and Saikumar, P., “Mitochondrial Remodeling Following Fission Inhibition by 15d-PGJ2 Involves Molecular Changes in Mitochondrial Fusion Protein OPA1”, Biochemical and biophysical research communications, vol. 399, pp. 548–554, 2010.[Abstract]


We showed earlier that 15 deoxy Δ12, 14 prostaglandin J2 (15d-PGJ2) inactivates Drp1 and induces mitochondrial fusion [1]. However, prolonged incubation of cells with 15d-PGJ2 resulted in remodeling of fused mitochondria into large swollen mitochondria with irregular cristae structure. While initial fusion of mitochondria by 15d-PGJ2 required the presence of both outer (Mfn1 and Mfn2) and inner (OPA1) mitochondrial membrane fusion proteins, later mitochondrial changes involved increased degradation of the fusion protein OPA1 and ubiquitination of newly synthesized OPA1 along with decreased expression of Mfn1 and Mfn2, which likely contributed to the loss of tubular rigidity, disorganization of cristae, and formation of large swollen degenerated dysfunctional mitochondria. Similar to inhibition of Drp1 by 15d-PGJ2, decreased expression of fission protein Drp1 by siRNA also resulted in the loss of fusion proteins. Prevention of 15d-PGJ2 induced mitochondrial elongation by thiol antioxidants prevented not only loss of OPA1 isoforms but also its ubiquitination. These findings provide novel insights into unforeseen complexity of molecular events that modulate mitochondrial plasticity.

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2010

Dr. Nandita Mishra, Kar, R., Singha, P. K., Venkatachalam, M. A., McEwen, D. G., and Saikumar, P., “Inhibition of Mitochondrial Division Through Covalent Modification of Drp1 Protein by 15 deoxy-Delta(12,14)-prostaglandin J2”, Biochemical and biophysical research communications, vol. 395, pp. 17–24, 2010.[Abstract]


Arachidonic acid derived endogenous electrophile 15d-PGJ2 has gained much attention in recent years due to its potent anti-proliferative and anti-inflammatory actions mediated through thiol modification of cysteine residues in its target proteins. Here, we show that 15d-PGJ2 at 1µM concentration converts normal mitochondria into large elongated and interconnected mitochondria through direct binding to mitochondrial fission protein Drp1 and partial inhibition of its GTPase activity. Mitochondrial elongation induced by 15d-PGJ2 is accompanied by increased assembly of Drp1 into large oligomeric complexes through plausible intermolecular interactions. The role of decreased GTPase activity of Drp1 in the formation of large oligomeric complexes is evident when Drp1 is incubated with a non-cleavable GTP analog, GTPγS or by a mutation that inactivated GTPase activity of Drp1 (K38A). The mutation of cysteine residue (Cys644) in the GTPase effector domain, a reported target for modification by reactive electrophiles, to alanine mimicked K38A mutation induced Drp1 oligomerization and mitochondrial elongation, suggesting the importance of cysteine in GED to regulate the GTPase activity and mitochondrial morphology. Interestingly, treatment of K38A and C644A mutants with 15d-PGJ2 resulted in super oligomerization of both mutant Drp1s indicating that 15d-PGJ2 may further stabilize Drp1 oligomers formed by loss of GTPase activity through covalent modification of middle domain cysteine residues. The present study documents for the first time the regulation of a mitochondrial fission activity by a prostaglandin, which will provide clues for understanding the pathological and physiological consequences of accumulation of reactive electrophiles during oxidative stress, inflammation and degeneration.

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Publication Type: Conference Proceedings

Year of Publication Title

2020

Anupama Binoy, Divya Nedungadi, Damu Sunilkumar, Dr. Bipin G. Nair, and Dr. Nandita Mishra, “Role of AIF in Plumbagin induced Paraptosis-A caspase independent cell death in MDA-MB-231 cancer cells”, 39th Annual Conference of Indian Association for Cancer Research (IACR-2020): “Leading the Fight against Cancer”, Rajiv Gandhi Centre for Biotechnology (RGCB), Trivandrum, Kerala, India, February 5-7. 2020.

2017

Divya Nedungadi, Megha Prasad, Anupama Binoy, Patra, G., Silva, J. D., Chatterjee, M., Jacob, A., Babu, A. S., Alexin, R., Dr. Bipin G. Nair, Dr. Sanjay Pal, and Dr. Nandita Mishra, “Anthelmintic properties of ginger compounds”, 27th Swadeshi Science Congress, November 7-9, 2017. Kerala, India, 2017.

2015

D. Nedungadi, ,, Pandurangan Nanjan, Nair, B. G., Dr. Asoke Banerji, and Dr. Nandita Mishra, “Caspase Independent Cell Death in Malignant Breast Cancer Cells MDA MB-231 by Chalcone Derivatives”, XXXIX All India Cell Biology Conference,IISER. Trivandrum, India, 2015.

2013

Dr. Nandita Mishra, Dr. Sanjay Pal, Nair, R. R., Krishna, A., Akhila Ajith, V, A., KS, D., Divya Nedungadi, and P, C., “Expression and refolding of recombinant staphylococcal amidases in E. coli”, Proceedings of International Conference on Biotechnology for Innovative Applications (Amrita BioQuest 2013). Elsevier Publications, p. 106, 2013.[Abstract]


Peptidoglycan (PG) hydrolases or autolysins are a group of enzymes which degrades bacterial cell wall at specific sites. Staphylococcus aureus produces two major PG hydrolases: major autolysin (Atl) and Aaa, a autolysin/adhesin protein. Amidase are surface-associated proteins that have both enzymatic and adhesive functions. AtlA is the most predominant autolysin in Staphylococcus aureus. The AltA protein only with an amidase domain and the two repeat sequences R1 and R2 is 62 kDa. Another autolysin/ adhesin present on the cell surface of S. aureus (Aaa) is a 35 kDa protein containing two direct LysM (lysine motif) repeats at the N-terminal and catalytic domain in the C-terminus. The amidase domain of the atlA gene with two repeat regions (amiE-R1,2) was cloned and expressed as N-terminal His-tag fusion protein in pQE30 (Qiagen) vector in E.coli M15 (Biswas et al., 2006). For the over-expression of His-tag Aaa; the aaa open reading frame was cloned in the pET28 (Novagen) vector designed to express proteins as fusions with a His6-tag at the N-terminus (Biswas R, 2006). We grew the E. coli M15 cells carrying the altA gene encoding the His-tagged protein in LB medium containing ampicillin to an OD600 nm of 0.5; expression was then induced with different concentrations of isopropyl β-D-thiogalactoside (IPTG). Cells were harvested by centrifugation, and lysed by freeze thaw method with the addition of lysozyme. Proteins were purified under denaturing conditions with 8M urea and analysed by 10 % SDS PAGE. In the same way, we also cultured the E. coli BL21 cells carrying the aaa gene but in LB medium with kanamycin instead of ampicillin. The optimal IPTG concentration for both was found to be 1mM. Isolation of insoluble and soluble fractions of recombinant protein from the bacterial cell lysate and their analysis confirmed the presence of both the recombinant proteins in the insoluble fraction possibly as inclusion bodies. Large scale purification of both the recombinant proteins was done using nickel (Ni-NTA) affinity chromatography. Maximum protein elution was found in 250mM imidazole buffer in both the cases. Refolding was optimized in refolding buffer containing 0.5mM reduced glutathione: 0.5mM oxidised glutathione (1:1) in 50mM Tris HCl pH-8.0. After refolding the bioactivity of both the proteins were assayed. Cell lysis activity of the refolded protein was observed to be more for 61KDa protein than the 35KDa protein. The refolded 61kDa amidase was found to be strongly binding to fibronectin found by affinity column chromatography.

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2011

Dr. Nandita Mishra, Kar, R., Singha, P. K., Venkatachalam, M. A., and Saikumar, P., “Ethacrynic acid induced mitochondrial fusion is mediated by Inhibition of fission protein Drp1”, XXXV All India Cell Biology Conference,16th – 18th Dec . NISER, Bhubaneswar, India, 2011.

2010

P. K. Singha, Dr. Nandita Mishra, Kar, R., Pandeswara, S., Venkatachalam, M. A., and Saikumar, P., “Targeting LC3-SQSTM1 (p62) signaling axis to treat apoptosis resistant and metastatic breast cancers”, American Association for Cancer Research Annual Meeting, April 17-21. Washington DC , 2010.

Publication Type: Patent

Year of Publication Title

2018

S. Bhuniya, Dr. Nandita Mishra, Velusamy, N., Anupama Binoy, Bobba, K. Naidu, and Divya Nedungadi, “Flourescent Exomarker Probes for Hydrogen Sulfide Detection”, U.S. Patent 15 / 956 , 4742018.[Abstract]


A fluorescence probe with mitochondrial targeting and two-photon property, its preparation method and application in detecting and tracking endogenous H2S in samples or living cells. The fluorescent probe is prepared by a four-step preparation method and demonstrates a UV-vis absorption increment λab=395 nm and ˜43 fold higher fluorescence intensity in the presence of H2S. The probe further demonstrates stability, selectivity for H2S over competing agents and sensitivity as low as 20 nm. A method of detecting endogenous H2S rapidly in the absence of any external stimulators is provided. Samples are contacted with the probe and the changes in fluorescence are monitored to detect H2S levels. The disclosed probe is non-toxic and suitable as a biomarker and therapeutic molecule in cancer and other diseases.

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