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Papers by Tushar Aggarwal

ABSTRACTPeroxynitrite (ONOO−), a redox-active species generated under nitrosative stress conditio... more ABSTRACTPeroxynitrite (ONOO−), a redox-active species generated under nitrosative stress conditions, can promote oxidative damage of lipids. Herein, we report the development and multifaceted utilization of a novel, activity-based phospholipid probe, PLP-ONOO–, for imaging lipid environments that are targeted by ONOO− in biomimetic and biological systems. Using PLP-ONOO– and the natural lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, we built synthetic protocell membranes in the form of giant vesicles (GVs). These GVs responded to ONOO− by lighting up and displayed excellent selectivity against other redox-active species, introducing an unprecedented function to biomimetic membrane architectures. Live HeLa cells containing PLP-ONOO– were investigated under cellular nitrosative stress induced by either external administration of peroxynitrite or endogenous stimulation with interferon-γ / lipopolysaccharide / phorbol myristate acetate. Both conditions enhanced the coumarin flu...

SSRN Electronic Journal, 2022

MicroRNA (miRNA)-based intercellular communication has been implicated in many functional and dys... more MicroRNA (miRNA)-based intercellular communication has been implicated in many functional and dysfunctional biological processes. This has raised interest in the potential use of miRNAs as biomarkers for diagnosis and prognosis. Though the list of clinically significant miRNA biomarkers is expanding, it remains challenging to adapt current chemical tools to investigate miRNAs in complex environments native to cells and tissues. We describe here a methodology for rapidly developing aptamer-based fluorescent biosensors that can specifically detect miRNAs in biologically relevant media (10-30% v/v), including medium collected from cultured HeLa cells, human serum, and human plasma. This methodology involves the semi-rational design of the hybridization between DNA oligonucleotides and the miRNA target to build a pool of potential aptamers, and the screening of this pool for high signal-to-background ratio and target specificity. The DNA oligonucleotides are readily available and require no chemical modification, rendering these chemical tools highly adaptable to any novel and niche miRNA target. Following this approach, we developed sensors that detect distinct oncogenic miRNA targets (miR-19b, miR-21, and miR-92a) at concentrations as low as 5 nM without amplification and are selective against single-nucleotide mutants. This work provides a systematic approach toward the development of miRNA biosensors that are easily accessible and can perform in biological environments with minimal sample handling.

The functionalization of material surfaces with biologically active molecules is crucial for enab... more The functionalization of material surfaces with biologically active molecules is crucial for enabling technologies in life sciences, biotechnology, and medicine. However, achieving biocompatibility and bioorthogonality with current synthetic methods remains a challenge. We report herein a novel surface functionalization method that proceeds chemoselectively and without a free transition metal catalyst. In this method, a coating is first formed via the tyrosinase-catalyzed putative polymerization of a tetrazine-containing catecholamine (DOPA-Tet). One or more types of molecule of interest containing trans-cyclooctene are then grafted onto the coating via tetrazine ligation. The entire process proceeds under physiological conditions and is suitable for grafting bioactive molecules with diverse functions and structural complexities. Utilizing this method, we functionalized material surfaces with enzymes (alkaline phosphatase, glucose oxidase, horseradish peroxidase), a cyclic peptide (...

ABSTRACTDNA polymerases have evolved to feature a highly conserved activity across the tree of li... more ABSTRACTDNA polymerases have evolved to feature a highly conserved activity across the tree of life: formation of, without exception, phosphodiester linkages that create the repeating sugarphosphate backbone of DNA. Can this linkage selectivity observed in nature be overcome by design to produce non-natural nucleic acids? Here, we report that structure-guided redesign of an archaeal DNA polymerase (9°N) enables a new polymerase activity that is undetectable in the wild type enzyme: catalyzing the formation of N3’→P5’ phosphoramidate linkages in the presence of 3’-amino-2’,3’-dideoxynucleoside 5’-triphosphate (3’-NH2-ddNTP) building blocks. Replacing a highly conserved metal-binding aspartate in the 9°N active site (Asp-404) with asparagine was key to the emergence of this unnatural enzyme activity. Molecular dynamics simulations provided insights into how a single substitution could enhance the productive positioning of the 3’-amino nucleophile in the active site. Further remodeling...

Volume 5 - 2020, Issue 8 - August

Prolene Hernia System is a tension free anterior inguinal hernia repair using a bilayered modific... more Prolene Hernia System is a tension free anterior inguinal hernia repair using a bilayered modification of inguinal hernia mesh. Inguinal hernia repair by Prolene is comparable with almost equal operating time, smaller incision and with a trend towards decreased complications rate and reduced rate of recurrences. This study was conducted to study and to compare the results of Prolene Hernia System and Lichtenstein Tension Free mesh hernioplasty with respect to its operative time, post-operative pain, intra/post-operative complications and total hospital stay. The effectiveness of PHS were compared to Lichtenstein tension free mesh hernias repair in patients presenting with uncomplicated Inguinal Hernia for elective surgery in the Surgery department of IGMC, Shimla. Duration of surgery was shorter in Prolene Hernia System group (p=0.04) than the Lichtenstein tension free mesh hernioplasty [32 vs 34 minutes], The mean pain intensity in present study was 2.9 in Lichtenstein tension free...

ABSTRACTIntercellular microRNA (miRNA)-based communication has been implicated in a wide array of... more ABSTRACTIntercellular microRNA (miRNA)-based communication has been implicated in a wide array of functional and dysfunctional biological processes. This has raised attention to the potential use of miRNAs as biomarkers for disease diagnosis and prognosis and produced interest in their detection. Though the list of clinically significant miRNA biomarkers is rapidly expanding, it remains challenging to adapt current tools to investigate new targets in biological environments. Systematic approaches for the rapid development of miRNA biosensors are valuable to reduce this disparity. We describe here a methodology for developing aptamer-based fluorescent biosensors that can specifically detect miRNAs in biological environments, including culture medium from HeLa cells, human serum, and human plasma. This methodology includes the semi-rational design of the hybridization between a pair of split DNA aptamer oligonucleotides and the miRNA target to build a pool of potential sensor designs,...

ABSTRACTPeroxynitrite (ONOO−), a redox-active species generated under nitrosative stress conditio... more ABSTRACTPeroxynitrite (ONOO−), a redox-active species generated under nitrosative stress conditions, can promote oxidative damage of lipids. Herein, we report the development and multifaceted utilization of a novel, activity-based phospholipid probe, PLP-ONOO–, for imaging lipid environments that are targeted by ONOO− in biomimetic and biological systems. Using PLP-ONOO– and the natural lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, we built synthetic protocell membranes in the form of giant vesicles (GVs). These GVs responded to ONOO− by lighting up and displayed excellent selectivity against other redox-active species, introducing an unprecedented function to biomimetic membrane architectures. Live HeLa cells containing PLP-ONOO– were investigated under cellular nitrosative stress induced by either external administration of peroxynitrite or endogenous stimulation with interferon-γ / lipopolysaccharide / phorbol myristate acetate. Both conditions enhanced the coumarin flu...

SSRN Electronic Journal, 2022

MicroRNA (miRNA)-based intercellular communication has been implicated in many functional and dys... more MicroRNA (miRNA)-based intercellular communication has been implicated in many functional and dysfunctional biological processes. This has raised interest in the potential use of miRNAs as biomarkers for diagnosis and prognosis. Though the list of clinically significant miRNA biomarkers is expanding, it remains challenging to adapt current chemical tools to investigate miRNAs in complex environments native to cells and tissues. We describe here a methodology for rapidly developing aptamer-based fluorescent biosensors that can specifically detect miRNAs in biologically relevant media (10-30% v/v), including medium collected from cultured HeLa cells, human serum, and human plasma. This methodology involves the semi-rational design of the hybridization between DNA oligonucleotides and the miRNA target to build a pool of potential aptamers, and the screening of this pool for high signal-to-background ratio and target specificity. The DNA oligonucleotides are readily available and require no chemical modification, rendering these chemical tools highly adaptable to any novel and niche miRNA target. Following this approach, we developed sensors that detect distinct oncogenic miRNA targets (miR-19b, miR-21, and miR-92a) at concentrations as low as 5 nM without amplification and are selective against single-nucleotide mutants. This work provides a systematic approach toward the development of miRNA biosensors that are easily accessible and can perform in biological environments with minimal sample handling.

The functionalization of material surfaces with biologically active molecules is crucial for enab... more The functionalization of material surfaces with biologically active molecules is crucial for enabling technologies in life sciences, biotechnology, and medicine. However, achieving biocompatibility and bioorthogonality with current synthetic methods remains a challenge. We report herein a novel surface functionalization method that proceeds chemoselectively and without a free transition metal catalyst. In this method, a coating is first formed via the tyrosinase-catalyzed putative polymerization of a tetrazine-containing catecholamine (DOPA-Tet). One or more types of molecule of interest containing trans-cyclooctene are then grafted onto the coating via tetrazine ligation. The entire process proceeds under physiological conditions and is suitable for grafting bioactive molecules with diverse functions and structural complexities. Utilizing this method, we functionalized material surfaces with enzymes (alkaline phosphatase, glucose oxidase, horseradish peroxidase), a cyclic peptide (...

ABSTRACTDNA polymerases have evolved to feature a highly conserved activity across the tree of li... more ABSTRACTDNA polymerases have evolved to feature a highly conserved activity across the tree of life: formation of, without exception, phosphodiester linkages that create the repeating sugarphosphate backbone of DNA. Can this linkage selectivity observed in nature be overcome by design to produce non-natural nucleic acids? Here, we report that structure-guided redesign of an archaeal DNA polymerase (9°N) enables a new polymerase activity that is undetectable in the wild type enzyme: catalyzing the formation of N3’→P5’ phosphoramidate linkages in the presence of 3’-amino-2’,3’-dideoxynucleoside 5’-triphosphate (3’-NH2-ddNTP) building blocks. Replacing a highly conserved metal-binding aspartate in the 9°N active site (Asp-404) with asparagine was key to the emergence of this unnatural enzyme activity. Molecular dynamics simulations provided insights into how a single substitution could enhance the productive positioning of the 3’-amino nucleophile in the active site. Further remodeling...

Volume 5 - 2020, Issue 8 - August

Prolene Hernia System is a tension free anterior inguinal hernia repair using a bilayered modific... more Prolene Hernia System is a tension free anterior inguinal hernia repair using a bilayered modification of inguinal hernia mesh. Inguinal hernia repair by Prolene is comparable with almost equal operating time, smaller incision and with a trend towards decreased complications rate and reduced rate of recurrences. This study was conducted to study and to compare the results of Prolene Hernia System and Lichtenstein Tension Free mesh hernioplasty with respect to its operative time, post-operative pain, intra/post-operative complications and total hospital stay. The effectiveness of PHS were compared to Lichtenstein tension free mesh hernias repair in patients presenting with uncomplicated Inguinal Hernia for elective surgery in the Surgery department of IGMC, Shimla. Duration of surgery was shorter in Prolene Hernia System group (p=0.04) than the Lichtenstein tension free mesh hernioplasty [32 vs 34 minutes], The mean pain intensity in present study was 2.9 in Lichtenstein tension free...

ABSTRACTIntercellular microRNA (miRNA)-based communication has been implicated in a wide array of... more ABSTRACTIntercellular microRNA (miRNA)-based communication has been implicated in a wide array of functional and dysfunctional biological processes. This has raised attention to the potential use of miRNAs as biomarkers for disease diagnosis and prognosis and produced interest in their detection. Though the list of clinically significant miRNA biomarkers is rapidly expanding, it remains challenging to adapt current tools to investigate new targets in biological environments. Systematic approaches for the rapid development of miRNA biosensors are valuable to reduce this disparity. We describe here a methodology for developing aptamer-based fluorescent biosensors that can specifically detect miRNAs in biological environments, including culture medium from HeLa cells, human serum, and human plasma. This methodology includes the semi-rational design of the hybridization between a pair of split DNA aptamer oligonucleotides and the miRNA target to build a pool of potential sensor designs,...