BITS Faculty Publications
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Item Acetylcholine structure-based small activatable fluorogenic probe for specific detection of acetylcholinesterase(ACS, 2023-05) Sidhu, Jagpreet SinghEarly detection of Alzheimer’s disease (AD) is important for taking proper measures against AD pathogenesis. Acetylcholinesterase (AChE) is widely reported to be associated with the pathogenicity of AD. Here, employing the “acetylcholine-mimic” approach, we designed and synthesized a new class of naphthalimide (Naph)-based fluorogenic probes for specific detection of AChE and avoiding interference of butyrylcholinesterase (BuChE), the pseudocholinesterase. We investigated the action of the probes on Electrophorus electricus AChE, and the native human brain AChE that we expressed in Escherichia coli and purified in the active form for the first time. The probe Naph-3 exhibited a substantial fluorescence enhancement with AChE and majorly avoided BuChE. Naph-3 successfully crossed the cell membrane of the Neuro-2a cells and fluoresced upon reaction with endogenous AChE. We further established that the probe could be effectively used for screening AChE inhibitors. Our study provides a new avenue for the specific detection of AChE, which can be extended to the diagnosis of AChE-related complications.Item Acetylcholine Structure-Based Small Activatable Fluorogenic Probe for Specific Detection of Acetylcholinesterase(ACS, 2023-05) Sidhu, Jagpreet SinghEarly detection of Alzheimer’s disease (AD) is important for taking proper measures against AD pathogenesis. Acetylcholinesterase (AChE) is widely reported to be associated with the pathogenicity of AD. Here, employing the “acetylcholine-mimic” approach, we designed and synthesized a new class of naphthalimide (Naph)-based fluorogenic probes for specific detection of AChE and avoiding interference of butyrylcholinesterase (BuChE), the pseudocholinesterase. We investigated the action of the probes on Electrophorus electricus AChE, and the native human brain AChE that we expressed in Escherichia coli and purified in the active form for the first time. The probe Naph-3 exhibited a substantial fluorescence enhancement with AChE and majorly avoided BuChE. Naph-3 successfully crossed the cell membrane of the Neuro-2a cells and fluoresced upon reaction with endogenous AChE. We further established that the probe could be effectively used for screening AChE inhibitors. Our study provides a new avenue for the specific detection of AChE, which can be extended to the diagnosis of AChE-related complications.Item Gold conjugated carbon dots nano assembly: FRET paired fluorescence probe for cysteine recognition(Elsevier, 2019-03) Sidhu, Jagpreet SinghThe detection and discrimination of Cys amino acid from numerous other related biomolecules has great importance in clinical field for diagnosis of various diseases. Herein, to detect the Cys, we embedded the carbon dots (CDs), gold, and naphthalimide (L1) into a single ratiometric fluorescence sensor assembly. Sensor assembly works on the principle of FRET mechanism between CDs and naphthalimide when CDs and L1 adhered on gold nanoparticles surface. Gold metal was turned into solid support by in situ reduction of HAuCl4 in the presence of CDs and L1. When the assembly was excited at 360 nm, emission maxima at 568 nm corresponded to naphthalimide emission was emerged that signifies the existence of a FRET between the CDs and naphthalimide fluorophores. With the addition of Cys, the FRET mechanism eliminated and the change in the fluorescence emission at two different wavelengths (450 nm and 568 nm) was recorded. The endogenous images of Cys was recorded by collecting the fluorescence images of HeLa cells under fluorescence microscope and also applied for the assay of Cys in blood serum. Cytotoxicity studies of CDs and sensor assembly were evaluated by performing the MTT assay.Item Synthesis of a 3,4-Disubstituted 1,8-Naphthalimide-Based DNA Intercalator for Direct Imaging of Legionella pneumophila(ACS, 2019-03) Sidhu, Jagpreet SinghThe development of organic molecules to target nucleic acid is an active area of research at the interface of chemistry and biochemistry, which involves DNA binding, nuclear imaging, and antitumor studies. These molecules bind with DNA through covalent interactions, electrostatic interactions, or intercalation. However, they are less permeable to membrane, and they have a significant cytotoxicity, which limits their application under in vivo conditions. In the present work, various mono- and disubstituted 1,8-naphthalimides-based derivatives (S-12, S-13, S-15, and S-21) have been synthesized and characterized through various spectroscopic techniques. Among these, 3-amino-4-bromo-1,8-naphthalimide (S-15) was found to have an attractive water solubility and act as a nuclear imaging agent. The spectroscopic absorption and emission data showed that S-15 has a strong affinity for salmon sperm DNA with a binding constant of 6.61 × 104 M–1, and the ratiometric fluorescence intensity (I489/I552) of S-15 has a linear relationship in the 0–50 μM range of DNA concentrations. It intercalates with DNA through the hydrophobic planar naphthalimide core as confirmed through cyclic voltammetry, circular dichroism, 1H NMR titration, and thermal denaturation studies. Positively charged amine groups also participate in H-bonding with the bases and backbone of DNA. The S-15 intercalator showed a large Stokes shift and photostability, which made it attractive for direct imaging of Legionella pneumophila, without the need for a prior membrane permeabilization.