Publications by authors named "Sourav Chowdhury"

20 Publications

  • Page 1 of 1

Development of antibacterial compounds that constrain evolutionary pathways to resistance.

Elife 2021 07 19;10. Epub 2021 Jul 19.

Department of Chemistry and Chemical Biology, Harvard University, Cambridge, United States.

Antibiotic resistance is a worldwide challenge. A potential approach to block resistance is to simultaneously inhibit WT and known escape variants of the target bacterial protein. Here, we applied an integrated computational and experimental approach to discover compounds that inhibit both WT and trimethoprim (TMP) resistant mutants of dihydrofolate reductase (DHFR). We identified a novel compound (CD15-3) that inhibits WT DHFR and its TMP resistant variants L28R, P21L and A26T with IC 50-75 µM against WT and TMP-resistant strains. Resistance to CD15-3 was dramatically delayed compared to TMP in in vitro evolution. Whole genome sequencing of CD15-3-resistant strains showed no mutations in the target folA locus. Rather, gene duplication of several efflux pumps gave rise to weak (about twofold increase in IC) resistance against CD15-3. Altogether, our results demonstrate the promise of strategy to develop evolution drugs - compounds which constrain evolutionary escape routes in pathogens.
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http://dx.doi.org/10.7554/eLife.64518DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8331180PMC
July 2021

Mixed Mott-Hubbard and charge transfer nature of 4H-SrMnOthin film on Si (100).

J Phys Condens Matter 2021 May 10;33(23). Epub 2021 May 10.

UGC-DAE Consortium for Scientific Research, Indore-452001, Madhya Pradesh, India.

Room temperature electronic structure of polycrystalline 4H-SrMnOthin film grown on Si (100) substrate has been studied using resonance photo emission spectroscopy and soft x-ray absorption spectroscopy measurements. Presence of charge transfer screen Mn 3final state along with the 3final state at the valence band edge of 4H-SrMnOthin film confirms that the ground state is strongly mixed between Mn 3and O 2states. The estimated equivalent values of on-site Coulomb interaction energy () and O 2to Mn 3- charge transfer energy (Δ) (≈ Δ ≈ 4.8 eV) from the combination of occupied and unoccupied spectra further confirm the intermediate Mott-Hubbard and charge transfer insulator nature of 4H-SrMnOfilm. Despite having similar Mn 4+ valence state in 4H-SrMnOand cubic SrMnO, 4H phase is observed to reveal much higher band gap ∼1.5 eV than the cubic phase (0.3 eV), which arises due to different MnOoctahedra environment.
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http://dx.doi.org/10.1088/1361-648X/abe8a3DOI Listing
May 2021

The metal cofactor zinc and interacting membranes modulate SOD1 conformation-aggregation landscape in an in vitro ALS model.

Elife 2021 Apr 7;10. Epub 2021 Apr 7.

Structural Biology & Bio-Informatics Division, CSIR-Indian Institute of Chemical Biology, Kolkata, India.

Aggregation of Cu-Zn superoxide dismutase (SOD1) is implicated in the motor neuron disease, amyotrophic lateral sclerosis (ALS). Although more than 140 disease mutations of SOD1 are available, their stability or aggregation behaviors in membrane environment are not correlated with disease pathophysiology. Here, we use multiple mutational variants of SOD1 to show that the absence of Zn, and not Cu, significantly impacts membrane attachment of SOD1 through two loop regions facilitating aggregation driven by lipid-induced conformational changes. These loop regions influence both the primary (through Cu intake) and the gain of function (through aggregation) of SOD1 presumably through a shared conformational landscape. Combining experimental and theoretical frameworks using representative ALS disease mutants, we develop a 'co-factor derived membrane association model' wherein mutational stress closer to the Zn (but not to the Cu) pocket is responsible for membrane association-mediated toxic aggregation and survival time scale after ALS diagnosis.
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http://dx.doi.org/10.7554/eLife.61453DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8087447PMC
April 2021

Evolutionary Analyses of Sequence and Structure Space Unravel the Structural Facets of SOD1.

Biomolecules 2019 12 4;9(12). Epub 2019 Dec 4.

Protein Folding and Dynamics Group, Structural Biology and Bio-informatics Division, CSIR-Indian Institute of Chemical Biology,4 Raja S.C.Mullick Road, Kolkata 700032, India.

Superoxide dismutase (SOD) is the primary enzyme of the cellular antioxidant defense cascade. Misfolding, concomitant oligomerization, and higher order aggregation of human cytosolic SOD are linked to amyotrophic lateral sclerosis (ALS). Although, with two metal ion cofactors SOD1 is extremely robust, the de-metallated apo form is intrinsically disordered. Since the rise of oxygen-based metabolism and antioxidant defense systems are evolutionary coupled, SOD is an interesting protein with a deep evolutionary history. We deployed statistical analysis of sequence space to decode evolutionarily co-varying residues in this protein. These were validated by applying graph theoretical modelling to understand the impact of the presence of metal ion co-factors in dictating the disordered (apo) to hidden disordered (wild-type SOD1) transition. Contact maps were generated for different variants, and the selected significant residues were mapped on separate structure networks. Sequence space analysis coupled with structure networks helped us to map the evolutionarily coupled co-varying patches in the SOD1 and its metal-depleted variants. In addition, using structure network analysis, the residues with a major impact on the internal dynamics of the protein structure were investigated. Our results reveal that the bulk of these evolutionarily co-varying residues are localized in the loop regions and positioned differentially depending upon the metal residence and concomitant steric restrictions of the loops.
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http://dx.doi.org/10.3390/biom9120826DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6995586PMC
December 2019

Ex vivo testing of air-cooled CW/modulated 30  W thulium fiber laser for lithotripsy.

Appl Opt 2019 Sep;58(25):6720-6724

A diode-pumped, air-cooled, all-fiber, quasi-continuous-wave thulium laser at an operating wavelength of 1.94 μm has been designed to study the performance of the laser parameter on the rate of fragmentation and its dependence on stone composition, fragmented particle size, as well as the retropulsion effect. The optimized laser cavity with an active fiber core/cladding diameter of 10/130 μm under a counter-propagating pump provides a stable laser power of 30 W at a slope efficiency of 50% and wall plug efficiency of 17%. The rate of fragmentation along with the retropulsion effect has been studied with human calcium oxalate monohydrate (COM) urinary stones (N=36) of different composition by using the designed laser and 200-μm-core low OH silica delivery fiber. The thulium fiber laser setting of 2.7 J pulse energy at the pulse rate of 10 Hz, pulse width of 90 ms, and peak power of 30 W is successful in breaking human COM stones in a controlled manner at a fragmentation rate of 0.8±0.4  mg/s, with almost uniform fragments of particle size less than 1.6 mm. During the stone fragmentation, the stone displacement (retropulsion effect) is less than 15 mm, even for the fragmented stone mass of 15±5  mg.
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http://dx.doi.org/10.1364/AO.58.006720DOI Listing
September 2019

Rogue waves in a linear cavity Yb-fiber laser through spectral filtering induced pulse instability.

Opt Lett 2019 May;44(9):2161-2164

In this Letter, experimental observation of dissipative rogue waves (DRWs) due to spectral filtering induced pulse instabilities in a mode-locked ytterbium (Yb) fiber laser has been presented. A semiconductor saturable absorber mirror was used to mode-lock the linear cavity laser and a chirped fiber Bragg grating (CFBG) was used for dispersion management, which also acted as a spectral filter and output coupler. Under stable conditions, the cavity delivered dispersion managed dissipative solitons of 447 fs duration and 0.69 nJ pulse energy at 10.19 MHz repetition rate with uniform intensity distribution over a long time span. As the spectral width increased with pump power, random intensity fluctuations were observed in the pulse train due to the filtering effect of the CFBG. Employing a dispersive Fourier transform by stretching the output pulse train in time allowed the existence of DRWs more than 4 times stronger than the significant wave height to be observed. Further increments of pump power led to a stable multi-pulsing state.
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http://dx.doi.org/10.1364/OL.44.002161DOI Listing
May 2019

Network mapping of the conformational heterogeneity of SOD1 by deploying statistical cluster analysis of FTIR spectra.

Cell Mol Life Sci 2019 Oct 22;76(20):4145-4154. Epub 2019 Apr 22.

Protein Folding and Dynamics Laboratory, Structural Biology and Bioinformatics Division, CSIR-Indian Institute of Chemical Biology, Kolkata, 700032, India.

A crucial contribution to the heterogeneity of the conformational landscape of a protein comes from the way an intermediate relates to another intermediate state in its journey from the unfolded to folded or misfolded form. Unfortunately, it is extremely hard to decode this relatedness in a quantifiable manner. Here, we developed an application of statistical cluster analyses to explore the conformational heterogeneity of a metalloenzyme, human cytosolic copper-zinc superoxide dismutase (SOD1), using the inputs from infrared spectroscopy. This study provides a quantifiable picture of how conformational information at one particular site (for example, the copper-binding pocket) is related to the information at the second site (for example, the zinc-binding pocket), and how this relatedness is transferred to the global conformational information of the protein. The distance outputs were used to quantitatively generate a network capturing the folding sub-stages of SOD1.
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http://dx.doi.org/10.1007/s00018-019-03108-2DOI Listing
October 2019

MYD88 and CXCR4 Mutation Profiling in Lymphoplasmacytic Lymphoma/Waldenstrom's Macroglobulinaemia.

Indian J Hematol Blood Transfus 2019 Jan 2;35(1):57-65. Epub 2018 Jul 2.

1Department of Laboratory Haematology and Molecular Genetics, Tata Medical Center, 14 MAR (EW), New Town, Rajarhat, Kolkata, 700156 India.

Recurrent mutations affecting MYD88 and CXCR4 gene nowadays form the basis for the diagnosis, risk stratification and use of inhibitors targeting these signalling pathways in LPL/WM which are rare B cell neoplasms. MYD88 L265P mutation analysis was performed on 33 cases of LPL/WM by AS-PCR (positivity-84.8%, n = 28/33) and by Sanger sequencing (positivity-39.3%, n = 13/33). We had only two cases with CXCR4 non-sense (NS) mutation (p.S338*) using Sanger sequencing. MYD88 (L265P) mutation detection by AS-PCR can form reliable biomarker for the diagnosis of LPL/WM in molecular labs. Although the cohort is small, still the CXCR4 mutation frequency in our study is low as compared to the published literature.
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http://dx.doi.org/10.1007/s12288-018-0978-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6369099PMC
January 2019

Understanding the evolutionary trend of intrinsically structural disorders in cancer relevant proteins as probed by Shannon entropy scoring and structure network analysis.

BMC Bioinformatics 2019 Feb 4;19(Suppl 13):549. Epub 2019 Feb 4.

Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts, 02138, USA.

Background: Malignant diseases have become a threat for health care system. A panoply of biological processes is involved as the cause of these diseases. In order to unveil the mechanistic details of these diseased states, we analyzed protein families relevant to these diseases.

Results: Our present study pivots around four apparently unrelated cancer types among which two are commonly occurring viz. Prostate Cancer, Breast Cancer and two relatively less frequent viz. Acute Lymphoblastic Leukemia and Lymphoma. Eight protein families were found to have implications for these cancer types. Our results strikingly reveal that some of the proteins with implications in the cancerous cellular states were showing the structural organization disparate from the signature of the family it constitutes. The sequences were further mapped onto respective structures and compared with the entropic profile. The structures reveal that entropic scores were able to reveal the inherent structural bias of these proteins with quantitative precision, otherwise unseen from other analysis. Subsequently, the betweenness centrality scoring of each residue from the structure network models was resorted to explore the changes in dependencies on residue owing to structural disorder.

Conclusion: These observations help to obtain the mechanistic changes resulting from the structural orchestration of protein structures. Finally, the hydropathy indexes were obtained to validate the sequence space observations using Shannon entropy and in-turn establishing the compatibility.
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http://dx.doi.org/10.1186/s12859-018-2552-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7394331PMC
February 2019

A novel tryptamine-appended rhodamine-based chemosensor for selective detection of Hg present in aqueous medium and its biological applications.

Anal Bioanal Chem 2019 Feb 9;411(6):1143-1157. Epub 2019 Jan 9.

Department of Chemistry, University of Calcutta, 92, A. P. C. Road, Kolkata, 700 009, India.

A novel rhodamine-tryptamine conjugate-based fluorescent and chromogenic chemosensor (RTS) for detection of Hg present in water was reported. After gradual addition of Hg in aqueous methanol solution of RTS, a strong orange fluorescence and deep-pink coloration were observed. The probe showed high selectivity towards Hg compared to other competitive metal ions. The 1:1 binding stoichiometry between RTS and Hg was established by Job's plot analysis and mass spectroscopy. Initial studies showed that the synthesized probe RTS possessed fair non-toxicity and effectively passed through cell walls of model cell systems, viz., human neuroblastoma (SHSY5Y) cells and cervical cells (HeLa) to detect intercellular Hg ions, signifying its utility in biological system. The limit of detection (LOD) was found to be 2.1 nM or 0.42 ppb by fluorescence titration. Additionally, the potential relevance of synthesized chemosensor for detecting Hg ions in environmental water samples has been demonstrated. Graphical abstract ᅟ.
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http://dx.doi.org/10.1007/s00216-018-1546-0DOI Listing
February 2019

The Role of Intestinal Fatty Acid Binding Proteins in Protecting Cells from Fatty Acid Induced Impairment of Mitochondrial Dynamics and Apoptosis.

Cell Physiol Biochem 2018 30;51(4):1658-1678. Epub 2018 Nov 30.

Protein Folding and Dynamics Laboratory, Structural Biology and Bioinformatics Division, CSIR-Indian Institute of Chemical Biology, Kolkata, India.

Background/aims: The conformation, folding and lipid binding properties of the intestinal fatty acid binding proteins (IFABP) have been extensively investigated. In contrast, the functional aspects of these proteins are not understood and matter of debates. In this study, we aim to address the deleterious effects of FA overload on cellular components, particularly mitochondria; and how IFABP helps in combating this stress by restoring the mitochondrial dynamics.

Methods: In the present study the functional aspect of IFABP under conditions of lipid stress was studied by a string of extensive in-cell studies; flow cytometry by fluorescence-activated cell sorting (FACS), confocal imaging, western blotting and quantitative real time PCR. We deployed ectopic expression of IFABP in rescuing cells under the condition of lipid stress. Again in order to unveil the mechanistic insights of functional traits, we arrayed extensive computational approaches by means of studying centrality calculations along with protein-protein association and ligand induced cluster dissociation. While addressing its functional importance, we used FCS and in-silico computational analyses, to show the structural distribution and the underlying mechanism of IFABP's action.

Results: Ectopic expression of IFABP in HeLa cells has been found to rescue mitochondrial morphological dynamics and restore membrane potential, partially preventing apoptotic damage induced by the increased FAs. These findings have been further validated in the functionally relevant intestinal Caco-2 cells, where the native expression of IFABP protects mitochondrial morphology from abrogation induced by FA overload. However, this native level expression is insufficient to protect against apoptotic cell death, which is rescued, at least partially in cells overexpressing IFABP. In addition, shRNA mediated IFABP knockdown in Caco-2 cells compromises mitochondrial dynamics and switches on intrinsic apoptotic pathways under FA-induced metabolic stress.

Conclusion: To summarize, the present study implicates functional significance of IFABP in controlling ligand-induced damage in mitochondrial dynamics and apoptosis.
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http://dx.doi.org/10.1159/000495672DOI Listing
January 2019

Protein Fibril-Templated Biomimetic Synthesis of Highly Fluorescent Gold Nanoclusters and Their Applications in Cysteine Sensing.

ACS Omega 2018 Jul 11;3(7):7703-7714. Epub 2018 Jul 11.

Protein Folding and Dynamics Laboratory, Structural Biology & Bio-Informatics Division, and Metabolic Disorder Laboratory, Cell Biology and Physiology Division, CSIR-Indian Institute of Chemical Biology, 4 Raja S. C. Mullick Road, Kolkata 700032, India.

Biomimetic synthesis of multifunctional fluorescent gold nanoclusters (Au NCs) is of great demand because of their ever-increasing applications. In this study, we have used self-assembled bovine serum albumin (BSA) amyloid-like nanofibers as the bioinspired scaffold for the synthesis of Au NCs. The amyloid fibril stabilized gold nanocluster (Fib-Au NC) has been found to have appreciable enhancement of fluorescence emission and a large 25 nm red shift in its emission maxima when compared to its monomeric protein counterpart (BSA-Au NC). The underlying mechanism accountable for the fluorescence behavior and its spectral shift has been thoroughly investigated by a combined use of spectroscopic and microscopic techniques. We have subsequently demonstrated the use of Fib-Au NCs for cysteine (Cys) sensing both in vitro and inside live cells. Additionally, cellular uptake and postpermeation effect of Fib-Au NCs have also been ascertained by detailed flow cytometry analysis, viability assay, and real-time apoptotic gene expression profiling.
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http://dx.doi.org/10.1021/acsomega.8b01033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6130899PMC
July 2018

Comparative Study of Toluidine Blue O and Methylene Blue Binding to Lysozyme and Their Inhibitory Effects on Protein Aggregation.

ACS Omega 2018 Mar 5;3(3):2588-2601. Epub 2018 Mar 5.

Organic and Medicinal Chemistry Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata 700 032, India.

A comparative binding interaction of toluidine blue O (TBO) and methylene blue (MB) with lysozyme was investigated by multifaceted biophysical approaches as well as from the aspects of in silico biophysics. The bindings were static, and it occurred via ground-state complex formation as confirmed from time-resolved fluorescence experiments. From steady-state fluorescence and anisotropy, binding constants were calculated, and it was found that TBO binds more effectively than MB. Synchronous fluorescence spectra revealed that binding of dyes to lysozyme causes polarity changes around the tryptophan (Trp) moiety, most likely at Trp 62 and 63. Calorimetric titration also depicts the higher binding affinity of TBO over MB, and the interactions were exothermic and entropy-driven. In silico studies revealed the potential binding pockets in lysozyme and the participation of residues Trp 62 and 63 in ligand binding. Furthermore, calculations of thermodynamic parameters from the theoretical docking studies were in compliance with experimental observations. Moreover, an inhibitory effect of these dyes to lysozyme fibrillogenesis was examined, and the morphology of the formed fibril was scanned by atomic force microscopy imaging. TBO was observed to exhibit higher potential in inhibiting the fibrillogenesis than MB, and this phenomenon stands out as a promising antiamyloid therapeutic strategy.
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http://dx.doi.org/10.1021/acsomega.7b01991DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6044680PMC
March 2018

Hybrid pumped gain-switched thulium fiber laser at a high repetition rate.

Appl Opt 2018 May;57(13):3546-3550

A gain-switched all-fiber thulium laser at 2 μm with high repetition rate has been demonstrated under a hybrid pumping scheme combined of a pulsed pump at 1.56 μm and CW pump at 793 nm. The in-band pulsed pump at 1.56 μm triggers the gain-switching pulses while the CW pump at 793 nm facilitates the energy storage. Therefore, the seed cavity delivers high energy pulses allowing the elimination of multistage amplification. Such hybrid pump configuration is effective for generating gain-switched pulses of high average power with better slope efficiency and pulse width of a few hundreds of a nanosecond. The optimized cavity under such hybrid pump configuration provides output power of 5.92 W from the gain-switched oscillator with slope efficiency of 60% and pulse width of 300 ns at a repetition rate of 344 kHz. Maximum pulse energy of 17.2 μJ and peak power of 53.9 W has been achieved at this repetition rate. Stable gain-switched pulses at reduced pump pulse energy have been achieved by the use of a CW pump at 793 nm. This novel pump configuration facilitates gain switch at higher repetition rates where enough pump pulse energy may not be available. The gain-switched laser also operates at 520 kHz and 1.3 MHz repetition rate by changing the 1.56 μm pulsed pump and cavity length.
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http://dx.doi.org/10.1364/AO.57.003546DOI Listing
May 2018

Diverse mode of operation of an all-normal-dispersion mode-locked fiber laser employing two nonlinear loop mirrors.

Appl Opt 2018 Feb;57(5):1225-1230

In this paper, we propose an all-normal-dispersion ytterbium-fiber laser with a novel ring cavity architecture having two nonlinear amplifying loop mirrors (NALM) as saturable absorbers, capable of delivering distinctly different pulses with adjustable features. By optimizing the loop lengths of the individual NALMs, the cavity can be operated to deliver Q-switched mode-locked (Q-ML) pulse bunches with adjustable repetition rates, mode-locked pulses in dissipative soliton resonance (DSR) regime or noise-like pulse (NLP) regime with tunable pulse width. The DSR pulses exhibit characteristic narrowband spectrum, while the NLPs exhibit large broadband spectrum. The operation regime of the laser can be controlled by adjusting the amplifier pump powers and the polarization controllers. To the best of the authors' knowledge, this is the first demonstration of a single mode-locked cavity where narrowband DSR pulses and broadband NLPs alongside Q-ML pulse bunches can be selectively generated by employing two NALMs.
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http://dx.doi.org/10.1364/AO.57.001225DOI Listing
February 2018

High repetition rate gain-switched 1.94  μm fiber laser pumped by 1.56  μm dissipative soliton resonance fiber laser.

Opt Lett 2017 Jul;42(13):2471-2474

A dissipative soliton resonance (DSR) mode-locked Er:Yb fiber laser has been used to pump a thulium fiber laser to generate gain-switched pulses at high repetition rates. Here 412 ns long DSR pulses with a center wavelength of around 1.56 μm at a repetition rate of 410 kHz have been fed to a thulium fiber laser, resulting in generation of gain-switched pulses at 1.94 μm. The minimum pulse width achieved was 256 ns with an average power of 4.6 W at 66% slope efficiency. Gain-switched pulses at 520 kHz and 750 kHz were generated through changing the pump pulse repetition rate by modifying the DSR cavity. To the best of our knowledge, this is the first demonstration of a high repetition rate gain-switched thulium fiber laser pumped by a DSR mode-locked fiber laser. As DSR pulses can be generated with high seed average power and energy independent of the operating wavelength regime as well as mode-locking technique, the proposed method can be applied to generate gain-switched pulses at high repetition rates and various wavelengths without the need of any optical or electrical modulators.
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http://dx.doi.org/10.1364/OL.42.002471DOI Listing
July 2017

Photoinduced Electron Transfer Switching Mechanism of a Naphthalimide Derivative with its Solvatochromic Behaviour: An Experimental and Theoretical Study with In Cell Investigations.

Chemistry 2017 Nov 26;23(65):16516-16524. Epub 2017 Oct 26.

Department of Chemistry, Jadavpur University, Kolkata, 700032, India.

The sole existence of a t-bone-shaped naphthalimide derivative [2-(2-aminoethyl)-1H-benzo[de]isoquinoline-1,3(2H)dione] (NAP), which gives rise to a photoinduced electron transfer (PET) mechanism, has been established using a combination of experimental and theoretical studies. In parallel an in vitro-in cell PET mechanism has also been shown. To understand the photophysics of NAP, solvent studies have been carried out in different solvents. In addition, theoretical calculations have been conducted to explain the spectroscopic properties through optimized structures. A "turn off" PET mechanism has also been observed in the presence of specific metal ions, namely, Cr , Fe and Hg among a series of metal ions. Theoretical studies reveal that NAP-Cr , NAP-Fe and NAP-Hg have their HOMO energy states lying in between a HOMO-LUMO energy state of the t-bone-type NAP molecule. On the contrary, the HOMO state of the other metal ion-NAP conjugate (NAP-M ) does not lie in between the HOMO-LUMO energy gap of the t-bone-type NAP molecule. Coupled with in vitro studies, in cell investigations reveal an enhancement of fluorescence intensity of NAP upon cytosolic metal sensing. Furthermore, a very high cell viability of NAP treated cells as tested by MTT assay and a fast permeation of the said compound as revealed by flow cytometry suggest NAP to be a potential candidate in metal sensing and bioimaging applications.
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http://dx.doi.org/10.1002/chem.201702414DOI Listing
November 2017

The Non-native Helical Intermediate State May Accumulate at Low pH in the Folding and Aggregation Landscape of the Intestinal Fatty Acid Binding Protein.

Biochemistry 2016 08 4;55(32):4457-68. Epub 2016 Aug 4.

Protein Folding and Dynamics Laboratory, Structural Biology and Bioinformatics Division, CSIR-Indian Institute of Chemical Biology , Kolkata 700032, India.

There has been widespread interest in studying early intermediate states and their roles in protein folding. The interest in intermediate states has been further emphasized in the recent literature because of their implications for protein aggregation. Unfortunately, direct kinetic characterization of intermediates has been difficult because of the limited time resolutions offered by the kinetic techniques and the heterogeneity of the folding and aggregation landscape. Even in equilibrium experiments, the characterization of intermediate states could be difficult because (a) their populations in equilibrium could be low and/or (b) they lack any specific biochemical or biophysical signatures for their identification. In this paper, we have used fluorescence correlation spectroscopy to study the nature of a low-pH intermediate state of the intestinal fatty acid binding protein, a small protein with predominantly β-sheet structure. Our results have shown that the pH 3 intermediate diffuses faster than the folded protein and has strong helix forming propensity. These behaviors support Lim's hypothesis according to which even an entirely β-sheet protein would form helical bundles at the early stage. Using dynamic light scattering and thioflavin T binding measurements, we have observed that the pH 3 intermediate is prone to aggregation. We believe that early helix formation is the result of a local effect, which originates from the interaction of the neighboring amino acids around the hydrophobic core residues. This early intermediate reorganizes subsequently, and this structural reorganization is initiated by the destabilizing interactions induced by the distant residues, unfavorable entropic costs, and steric constraints of the hydrophobic side chains. Mutational analyses show further that the increase in the hydrophobicity in the hydrophobic core region increases the population of the α-helical intermediate, enhancing the aggregation propensity of the protein, while an identical change, distant from the hydrophobic core, does not show any effect. This study re-emphasizes an overlap between the folding and aggregation landscape of a protein, where the fine-tuning between the local and global effects may be important for the protein to fold efficiently or to aggregate.
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http://dx.doi.org/10.1021/acs.biochem.6b00390DOI Listing
August 2016

Anencephaly and gastroschisis in the same foetus: a rarity.

J Indian Med Assoc 2012 Jul;110(7):503

Department of Obstetrics and Gynaecology, RG Kar Medical College and Hospital, Kolkata 700004.

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July 2012
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