Education and Training

Postdoctoral Training. (2006-2011: DNA Repair)

Dept. of Biochemistry and Molecular Biology, University of Texas Medical Branch (UTMB), Galveston, TX, USA

Ph.D. (2001-2006: Life Sciences)

School of Life Sciences, Jawaharlal Nehru University (JNU), New Delhi, India

M.Phil. (1998-2000: Biophysics)

Dept. of Biophysics, National Institute of Mental Health and Neurosciences (NIMHANS), Bangalore, India

M.Sc. (1996-1998: Biophysics)

Dr. Babasaheb Ambedkar Maratwada University, Aurangabad, Maharashtra, India

B.Sc. (1993-1996: Microbiology)

Osmania University, Hyderabad, Andhra Pradesh, India

Research training/fellowships

A. Positions and Honors

Positions Held

2012 -

Assistant Professor, Center for Biosciences, Central University of Punjab, Bathinda, Punjab, India

2011-12

Assistant Professor (Non-Tenure Research), Dept. of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, Texas, USA

2007-11

Postdoctoral Research Associate, Dept. of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, Texas, USA

2006-07

Postdoctoral Research Fellow, Dept. of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, Texas, USA

2003-05

Senior Research Fellow, ICMR project, School of Life Sciences, Jawaharlal Nehru University, New Delhi, India

2002-03

Senior Research Fellow, UGC-Excellence project, School of Life Sciences, Jawaharlal Nehru University, New Delhi, India

2001-02

Lecturer in Biophysics, Dept. of Physiology, S. V. S. Medical College, Mahabubnagar, India

2000-01

Lecturer in Biophysics, Dept. of Physiology, M. R. Medical College, Gulbarga, India

Honors

2011

Postdoctoral travel award to attend the American Society for Biochemistry and Molecular Biology (ASBMB) 2011 Annual Meeting, held in conjunction with Experimental Biology, in Washington, DC, April 8-13, 2011.

2010

Young Investigator travel award from the Alzheimer’s Drug Discovery Foundation (ADDF), USA (not accepted)

2009

Young Investigator award from the Alzheimer’s Drug Discovery Foundation (ADDF), USA

2004

Travel award for attending the International Brain Research Organization (IBRO) Advanced School on Neuroscience - “Receptors, Channels, Messengers” at Yalta, Ukraine, from CSIR, New Delhi, India

2002-2005

ICMR Senior Research Fellowship, Indian Council for Medical Research, New Delhi, India

1998-2000

NIMHANS Fellowship, National Institute of Mental Health and Neuro Sciences, Bangalore, India

Other Experience and Professional Memberships

2012-till date

Society for Neuroscience, Galveston Chapter, USA

2011-till date

Alzheimer’s Association, USA

2011-till date

Reviewed manuscripts for the “Journal of Developmental Neuroscience”

2010

Reviewed manuscripts for the Journal “Mutation Research”

2003-04

Indian Biophysical Society

B. Grant Support

Mantha AK (PI): APE1/Ref-1’s Dual Functions Countering Beta Amyloid Induced Genotoxicity. Alzheimer’s Association, USA, $99,990 for 2 years: 2011-2013.

Summary:

Human genome is constantly damaged due to intrinsic and extrinsic genotoxic agents that contribute towards oxidative DNA base damage. These oxidative DNA base lesions often mutagenic and toxic are repaired via the base excision repair pathway (BER) in which the abasic (AP) endonuclease, APE1, a multifunctional essential protein, plays a central role. APE1 also functions as a transcriptional co-regulator reductively activating several transcription factors (TFs, e.g., AP-1, NF-kB, and named Ref-1). Amyloid beta (Abeta) protein deposition in Alzheimer’s disease (AD) brains induces oxidative damage in neuronal genomes causing their death. APE1/Ref-1’s protective mechanism against Abeta-induced damage is poorly understood. I found Abeta(25-35) peptide treated rat PC12 cells accumulated AP sites, single-strand breaks (SSBs) and had activated poly [ADP-ribose] polymerase 1 (PARP-1), a SSB-sensor. Further, Abeta enhanced SSB-repair activity in PC12 nuclear extracts. In addition, APE1 over-expression decreased Abeta-induced reactive oxygen species (ROS). Abeta-stress also activated binding of AP-1 and NF-kB to their cis elements, which may be associated with uncontrolled gene expression. APE1-FLAG immunocomplexes of human SH-SY5Y neuroblastoma cells contained ADO (cysteamine [2-aminoethanethiol] dioxygenase) identified by 2D-proteomic approach. Abeta further enhanced ADO’s association with APE1, and furthermore, siRNA-mediated combined down-regulation of ADO and APE1 sensitized SH-SY5Y cells to Abeta-stress, whose physiological significance is unknown. Our preliminary studies along with the limited available information warrants to us perform in depth analysis to understand the role of APE1-dependent BER/SSBR-pathway to estimate the damage response of Abeta-stress, in defining the rate limiting enzyme(s). Further, characterization of ADO’s novel function as a “thiol exchanger” will establish that physiological and functional association of ADO with APE1 is for maintaining REDOX homeostasis. I will use cultured human SH-SY5Y neuroblastoma cells, and terminally differentiated neurons (retinoic acid and nerve grow factor treated SH-SY5Y cells) as a model to study AD-phenotype. Upon successful completion, the proposed studies may help define the neuroprotection rendered by APE1/Ref-1 utilizing its dual functions countering Abeta-induced genotoxicity.

C. Peer-reviewed and International Publications

1

Sengupta S, Chattopadhyay R, Mantha AK, Mitra S, and Bhakat KK. Regulation of mouse renin gene by AP-endonuclease 1(APE1/Ref-1) via recruitment of HDAC1 co-repressor complex. J. of Hypertension. 2012. (IF): 3.98

2

Hegde ML, Mantha AK, Hazra TK, Mitra S, and Szczesny B. Oxidative genome damage and its repair: Implications in aging and neurodegenerative diseases. Mechanism of Aging and Development. 2012: (IF): 4.85

3

#Mantha AK, Dhiman M, Perez-Polo JR and Mitra S. Proteomic Analysis Unravels APE1/Ref-1 Regulation of Beta Amyloid-Induced Neurotoxicity. J. of Neuroscience Research. 2012. (IF): 2.99 # Corresponding Author

4

Mantha AK*, Oezguen N*, Mitra S and Braun W. Additional evidence for the “moving ion” mechanism in endonuclease function of APE1. Bioinformation. 2011: 7(4):184-190. *Equal Authors. (IF): 1.15

5

Sengupta S, Mantha AK, Mitra S and Bhakat KK. Human AP-endonuclease (APE1/Ref-1) and its acetylation regulate YB-1/p300 recruitment and RNA polymerase II loading in the drug induced activation of multidrug resistance gene MDR1. Oncogene. 2010: 30(4):482-93. (IF): 7.41

6

Barnes T, Kim WC, Mantha AK, Kim SE, Izumi T, Mitra S, Lee CH. Identification of apurinic/apyrimidinic endonuclease APE1 as the endoribonuclease that cleaves c-myc mRNA. Nucleic Acids Research. 2009: 37(12):3946-58.

7

Bhakat KK, Mantha AK and Mitra S. Transcriptional Regulatory Functions of Mammalian AP-endonuclease (APE1/Ref-1), an Essential Multifunctional Protein. Antioxidant Redox Signaling. 2009; 11(3): 1-17. (IF): 8.2

8

Mantha AK, Oezguen N, Izumi T, Braun W and Mitra S. Unusual role of a cysteine residue in substrate binding and activity of human AP-endonuclease1. Journal of Molecular Biology. 2008; 379(1):28-37. (IF): 4.0.

9

Dike A, Chandrashekaran IR, Mantha AK, Baquer NZ and Cowsik SM. Pharmacophore Pattern Identification of Tachykinin Receptor Selective Peptide Agonists: Implications in Receptor Selectivity. American Journal of Biochemistry & Biotechnology. 2007; 3 (4): 180-186, (Review).

10

Bhakat KK, Chattopadhyay R, Tadahide I, Mantha AK, and Mitra S. Acetylated APE1 is a repressor in calcium-mediated down regulation of the human renin gene. Circulation. 2006; 114 (18): 129 (abstract). (IF): 14.59

11

Mantha AK, Moorthy K, Cowsik SM and Baquer NZ. Membrane associated functions of neurokinin B (NKB) on A? (25-35) induced toxicity in aging rat brain synaptosomes. Biogerontology. 2006 b; 7 (1): 19-33. (IF): 3.41

12

Mantha AK, Moorthy K, Cowsik SM and Baquer NZ. Neuroprotective role of neurokinin B (NKB) on ? - amyloid (25-35) induced toxicity in aging rat brain synaptosomes: Involvement in oxidative stress and excitotoxicity. Biogerontology. 2006 a; 7 (1): 1-17.

13

Moorthy K, Yadav UCS, Siddiqui MR, Mantha AK, Cowsik SM, Sharma D, Basir SF and Baquer NZ. Effect of hormone replacement therapy in normalizing age related neuronal markers in different age groups of naturally menopausal rats. Biogerontology. 2005; 6(5): 345-356. (IF): 3.41

14

Mantha AK, Chandrashekar IR, Baquer NZ and Cowsik SM. Three-dimensional structure of the mammalian tachykinin peptide neurokinin B bound to lipid micelles. Journal of Biomolecular Structure and Dynamics. 2004; 22(2): 137-148. (IF): 4.98

15

Moorthy K, Yadav UC, Mantha AK, Cowsik SM, Sharma D, Basir SF and Baquer NZ. Estradiol and Progesterone treatments change the lipid profile in naturally menopausal rats from different age groups. Biogerontology. 2004; 5 (6) 411-419. (IF): 3.41

16

Chandrashekar IR, Mantha AK, Dike A and Cowsik SM. Three-dimensional structure of lipid induced NK-2 selective tachykinin agonists. Journal of Biomolecular Structure and Dynamics. 2003; 20 (6) (abstract). (IF): 4.98

D. Posters Presented at International and National Levels

1

Sengupta S, Mantha AK, Mitra S and Bhakat KK. APE1 and its acetylation regulate YB-1/p300 complex formation and RNA pol II loading in the drug induced activation of MDR1 gene. (1st prize) at CANCER CENTER DAY, May 18th 2010 at UTMB, Galveston, TX.

2

Mantha AK, Hegde ML, Hazra TK, and Mitra S. Repair of Radiation-Induced Genome Damage: Collaboration between Base Excision and Non-homologous End Joining Repair. Atlanta May 15th to 19th 2010. (Oral presentation).

3

Tsutakawa S, Mantha AK, Ivanov I, Mol CD, Arvai A, Holton J, Frankel K, Cooper P, Cunningham R, Miitra S, and Tainer J. Common structural basis for apurinic/apyrimidinic endonuclease catalytic activity. March 2010, at SBDR meeting, Berkely, CA, USA.

4

Mantha AK, Dineley KT, Perez-Polo JR and Mitra S. Neuroprotective Role of APE1/Ref-1 in Amyloid Beta Mediated Genotoxicity in Neuronal Precursor PC12 Cells. November 12, 2009 at UTMB, Galveston, TX, USA.

5

Mantha AK, Oezguen N, Mitra S and Braun W. Additional Evidence For The “Moving Ion” Mechanism In Endonuclease Function of APE1/Ref-1. November, 2009 at Houston, TX, USA.

6

Mantha AK, Oezguen N, Izumi T, Bhakat KK, Mitra S and Braun W. Does the APE1 Double Mutant E96Q, D210N Bind Mg2+ in the Active Site? February 19 to 24, 2009 at Galveston, TX, USA.

7

Mantha AK, Bhakat KK, Oezguen N, Izumi T, Braun W and Mitra S. Exploring potential link between seven Cys residues of HAPE1’s repair and redox functions: a multi functional DNA repair enzyme. May 16 to 17, 2008 at Galveston, TX, USA.

8

Oezguen N, Mantha AK, Bhakat KK, Izumi T, Mitra S and Braun W. Influence of a distant Ser residue on the active site over 16Å: Unusual affect of C99S APE1 on endonuclease activity or repair function, May 16 to 17, 2008 at Galveston, TX, USA.

9

Tsutakawa SE, Hura GL, Mol CD, Izumi T, Arvai A, Mantha AK, Ivanov I, Mitra S, Tainer JA, Cooper PK. Structural analysis of the regulatory N-terminal domain of apurinic/apyrimidinic endonuclease (APE1). Joint EU-USA DNA repair workshop. April 11 to 14, 2007 at Berkeley, USA.

10

Mantha AK, Chattopadhyay R, Bhakat KK and Mitra S. Redox-regulating Cys residues in hAPE1. “Mammalian DNA Repair” Gorden research conference, February 4 to 9, 2007 at Ventura, CA, USA.

11

Bhakat KK, Chattopadhyay R, Izumi T, Mantha AK, and Mitra S. Acetylated APE1 is a repressor in calcium-mediated downregulation of the human renin gene. American Heart Association Scientific Section, November 12 to 15, 2006 at Chicago, USA.

12

Dike A, Chandrashekar IR, Mantha AK, Baquer NZ and Cowsik SM. Lipid induced conformation of the tachykinin neuropeptides Scyliorhinin I and Uperolein “SMASH 2005” on small molecule NMR, September 25 to 28, 2005 at Verona, Italy.

13

Mantha AK, Chandrashekar IR, Dike A, Moorthy K, Baquer NZ and Cowsik SM. Role of Neurokinin B and Ab-protein fragment 25-35 on aging rat brain synaptosomes. The XXIst International Conference on Magnetic Resonance in Biological Systems, January 16 to 21, 2005 at Hyderabad, India.

14

Chandrashekar IR, Mantha AK, Dike A and Cowsik SM. Structural features of Neurokinin-2 receptor selective peptide agonists and their role in receptor selectivity. The XXIst International Conference on Magnetic Resonance in Biological Systems, January 16 to 21, 2005 at Hyderabad, India.

15

Dike A, Chandrashekar IR, Mantha AK and Cowsik SM. Structural characterization of Scyliorhinin I: A dual NK1/NK2 receptor agonist. The XXIst International Conference on Magnetic Resonance in Biological Systems, January 16 to 21, 2005 at Hyderabad, India.

16

Mantha AK, Chandrashekar IR, Moorthy K, Baquer NZ and Cowsik SM. Tachykinin NK-3 Receptor and its Agonist Neurokinin B (NKB) Interactions: Shedding Light on Aging Brain Functions. IBRO Advanced School on Neuroscience “Receptors, Channels, Messengers”, September 16 to 28, 2004 at Yalta, Ukraine.

17

Chandrashekar IR, Mantha AK, Dike A and Cowsik SM. Lipid induced structure of Neurokinin-2 selective agonists. BaCaTeC Summer School on “NMR of Biomolecular complexes”, October 3 to 8, 2004 at Bayreuth, Germany.

18

Mantha AK, Chandrashekar IR, Dike A, Baquer NZ and Cowsik SM. Three-dimensional structure of the mammalian tachykinin peptide neurokinin B bound to lipid micelles: A NK3 receptor agonist of mammalian origin involved in neuroprotection and Aging. NMRS 2004 [National Magnetic Resonance Society] Symposium on NMR Drug Design & Bioinformatics, February 17 to 20, 2004 at Kolkatta, India.

19

Chandrashekar IR, Mantha AK, Dike A and Cowsik SM. Tachykinin neuropeptides: Membrane induced structure and receptor selectivity. Biosparks, February 14 to 15, 2004 at School of Life Sciences, JNU, New Delhi, India.

20

Moorthy K, Mantha AK, Basir SF and Baquer NZ. Neuroprotective role of Oestrogen and Progesterone in aging female rats. ACBICON, NIMHANS, January 19 to 22, 2004 at Bangalore, India.

21

Chandrashekar IR, Mantha AK, Dike A and Cowsik SM. Three dimensional structure of lipid induced NK-2 selective tachykinin agonists. 13th Conversation, June 17 to 21, 2003 at Albany, USA.

22

Mantha AK, Chandrashekar IR, Dike A, Baquer NZ and Cowsik SM. Lipid induced conformation of the mammalian tachykinin peptide neurokinin B. BIOSPARKS, School of Life Sciences, Jawaharlal Nehru University, March 31 to April 1, 2003 at New Delhi, India.

23

Vidyasagar S, Mantha AK and Kulkarni SG. Evaluation of radiation exposure doses from television. National Symposium on Radiation and Molecular Biophysics, Bhabha Atomic Research Centre (BARC), January 21 to 24, 1998 at Mumbai, India.

E. Area of Research and Research Perspectives

My research interest underscores examining various DNA repair and regulatory proteins to understand the molecular mechanisms and pathways through which the processes associated with onset of aging, age related, and neurodegenerative disorders in developing potential therapeutic targets. I am currently working on regulatory and repair functions of AP-endonuclease (APE1) in mammalian genomes. APE1 is the main apurinic/apyrimidinic endonuclease in eukaryotic cells playing a central role in the Base Excision repair (BER) pathway of all DNA lesions (uracil, alkylated and oxidized and abasic sites) including single-strand breaks and has role in co-transcriptional activation of TFs such as AP-1, NF-?B, TP53 and HIF1 a, and named redox effecter factor-1 (Ref-1). APE1/Ref-1 is a vital protein with its biological activities located in two functionally distinct domains. The N-terminus, containing the nuclear localization signal (NLS) region, is principally devoted to the redox activity and protein-protein interactions (my unpublished data), while the C-terminus exerts the enzymatic activity on the AP sites of DNA. The N-terminal also has post-translational modification “ACETYLATION”-sites at K6/K7, whose role in repair function of APE1/Ref-1 is still unclear. Over all APE1/Ref-1 contains 29 Lys residues in its 318-aa-long polypeptide, most of which are likely present on the APE1/Ref-1 surface and accessible by the ubiquitin ligase components. This ubiquitination found to be observed inside the N-terminal aa K24, K25, and K27 of APE1/Ref-1, which are completely conserved among mammals. In addition, these aa are also reported to be acetylated, and this N-terminal segment of APE1/Ref-1 acts as a device for fine regulation of protein catalytic activity on abasic DNA and co-ordination role in BER-pathway.

My own interest lies in elucidating the neuro-protective role of APE1/Ref-1 in neurological disorders e.g., Alzheimer’s disease (AD) and Parkinson’s disease (PD). Due to its multi functional nature; it is proving to be a critical target in neuronal cancer therapy for “Glioblastoma”. APE1/Ref-1’s role as a neuro-protector is an unexplored field of neurodegenerative disorders. Furthermore, the expression of APE1/Ref-1 and its sub-cellular localization is still poorly investigated in case of AD and PD. My research for the next 4-5 years mainly focused to understand and elucidate the molecular mechanisms of genotoxicity associated with amyloid beta (Aß) protein deposits in human brain tissue and role of APE1/Ref-1 as a potential candidate for therapeutic intervention in AD using neuronal cell lines (PC12 and SH-SY5Y) and transgenic mouse (Tg 2576 APP) as model systems.

Protein phosphorylation, another form of post-translational modification in APE1/Ref-1 reported to be observed on aa Thr233 by CDK5 in neurons of AD and PD brain tissues, further this phosphorylation results in decreased endonuclease function and pathogenesis of AD. Recently Dr Izumi’s group at LSU showed a cross talk between APE1/Ref-1’s monoubiquitination, mimicry of phosphorylation at aa Thr233 by CDK5. This might be a delicate balance of ubiquitination and phosphorylation activities that alter the gene regulatory function of APE1/Ref-1. In addition S-Nitosylation on Cys residues on crucial enzymes and proteins affecting neuronal cell’s ability to combat oxidative stress are known to progressive neurological and neurodegenerative disorders. It is yet to be studied in the case of APE1/Ref-1, my preliminary studies suggest that more than one Cys residue of APE1/Ref-1 are likely to be S-Nitrosylated upon Amyloid stress in experimental culture human SH-SY5Y cells, whose significance is one of my future research endeavors.

In addition, my research interest also shared by a small group of bioactive peptides known as Tachykinins (TKs); which demonstrate functional significance associated with their structural aspects. The mammalian TK peptides interact through G-protein coupled receptors (GPCRs). TKs have been identified and shown to be involved in a wide range of actions involving physiological and pathological conditions. The 25-35 aa peptide of Amyloid (1-42) shows sequence similarity with neurokinin B (NKB), substance P (SP) and neuronkinin A (NKA) of TK peptides and acts as agonist for NK- receptor functions. Earlier I have shown that (my doctoral studies from JNU, New Delhi), NKB a decapeptide of neurokinin-3 (NK-3) receptor agonist, acts as a neuro-protector against Ab-toxicity in aging rat brain synaptosomes. The detailed structure-function interactions between NKB and Ab, further might have significant insight into the mechanism of drug target development and which may be the potential site of interaction between the two peptides in case of neuroprotection, which is another area of my research interest for next couple of years.

Majorly my research utilizes cloning and purification of recombinant proteins, utilization of synthetic peptides, oligonucleotides, and oxidatively damaged DNA bases to mimic in vivo damage and repair processes in a test tube in laboratory conditions. I employ quantitative bio-physical techniques such as fluorescence spectroscopy (fluorescence anisotropy, FAN) and circular dichroism (CD). Biochemical techniques such as limited proteolytic digestion for identification of interacting domains and hot spots (critical amino acids) on critical proteins of interest along with Far-Western analysis, in vitro interaction studies by GST- as well as other affinity tagged pull downs from both recombinant as well as from both tissue and cultured cell extracts, and imaging techniques like confocal microscopy associated with immuno-histochemistry.

F. Past Research/Projects Completed

a. Doctoral Dissertation: “The role of neurokinin B (NKB) and amyloid beta protein fragment (25-35) on molecular and biochemical correlates in aging brain functions”.

: Tachykinins (TKs) comprise a group of bioactive peptides with functional significance associated with their structural aspects. The mammalian TK peptides interact through G-protein coupled receptors (GPCRs). TKs have been identified and shown to be involved in a wide range of actions involving physiological and pathological conditions. Amyloid beta protein is the main causative agent that is involved with Alzheimer disease (AD). The 25-35 amino acid peptide of amyloid (1-42) shows sequence similarity with neurokinin B (NKB), substance P (SP) and neuronkinin A (NKA) of TK peptides and acts as agonist for NK- receptor functions. In this study I have shown that, Neurokinin B (NKB) a decapeptide of neurokinin-3 (NK-3) receptor agonist, has neuroprotective role in aging rat brain associated with its structural and functional aspects.

The study involved techniques like, Circular Dichroism (CD), Fluorescence, and 2D-NMR spectroscopic techniques for structure determination of NKB. NMR studies indicated that NK-3 selective agonist NKB adopts a helical conformation in membrane mimetic DPC micelles (Mantha el al., 2004); a random coil conformation in aqueous medium further confirmed our hypothesis that NKB attains bioactive conformation when it interacts with the membrane for receptor mediated functions. Secondary structure prediction using CD suggested that Ca2+ ions do have additional structural constraints on NKB. Interaction of NKB with Ca2+ might have pivotal role in the bioactive conformation of associated with many of the physiological and pathological functions. Amyloid mediated cellular functions were studied by using isolated rat brain synaptosomes of different age groups. Enzyme activities [Acetylcholine esterase, Na+ - K+ ATPase, Monoamine oxidase (MAO), and Superoxide dismutase (SOD)] were measured in presence of these peptides singly or in combination. Ca2+ as a neuro-functional hallmark and its influence by these peptides were studied by Fura 2. Quantitative estimation of NKB receptor: NK-3 at different stages of age was done by Western blot analysis. It was summarized that NKB acts against amyloid mediated enzyme functions. Taking into consideration that free radicals, neurotransmitters, calcium and energy imbalance have important role in aging, it was concluded that NKB may delay/retard/protect against aging processes in the brain tissue by checking various enzyme functions (Mantha et al., 2006 a & b). Structure-function interactions between NKB and Ab (25-35), further might have significant insight into the mechanism of drug target development and which may be the potential site of interaction between the two peptides in case of neuroprotection.

b. M.Phil Dissertation: “Ca2+ Induced structural and functional changes in isolated rat brain mitochondria”.

In physiological and pathological conditions intracytoplasmic concentrations of calcium [Ca2+]i dramatically increases. This increase in intra cytoplasmic Ca2+ makes mitochondria to accumulate the excess of Ca2+ from cytosol leading to the mitochondrial Ca2+ overload, and the consequence are depletion in ATP synthesis, oxidative stress and production of excessive free radicals. What exactly is happening to mitochondria in case of mitochondrial Ca2+ overload is not clear. We have investigated the effect of Ca2+ on structural and functional aspects of mitochondria. The structural perturbations were studied by steady state anisotropy, and (a) measuring succinate dehydrogenase activity, and (b) measuring mitochondrial membrane potential studied the functional changes. In presence of Ca2+ these parameters changed dramatically suggesting that, these alterations might be the potential pathways for the mitochondrial dysfunction in case of mitochondrial Ca2+ overload in different neurological disorders. Ultra centrifugation, VIS/UV Spectroscopy, and Fluorescence spectroscopy were used for the above study.