Tricking enzymes in living cells: a mechanism-based strategy for design of DNA topoisomerase biosensors

AbstractMost activity-based molecular probes are designed to target enzymes that catalyze the breaking of chemical bonds and the conversion of a unimolecular substrate into bimolecular products. However, DNA topoisomerases are a class of enzymes that alter DNA topology without producing any molecular segments during catalysis, which hinders the development of practical methods for diagnosing these key biomarkers in living cells. Here, we established a new strategy for the effective sensing of the expression levels and catalytic activities of topoisomerases in cell-free systems and human cells. Using our newly designed biosensors, we tricked DNA topoisomerases within their catalytic cycles to switch on fluorescence and resume new rounds of catalysis. Considering that human topoisomerases have been widely recognized as biomarkers for multiple cancers and identified as promising targets for several anticancer drugs, we believe that these DNA-based biosensors and our design strategy would greatly benefit the future development of clinical tools for cancer diagnosis and treatment. Graphical Abstract

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Tricking Enzymes In Living Cells: A Mechanism-Based Strategy For Design of DNA Topoisomerase Biosensors

10.21203/rs.3.rs-720390/v1 ◽

2021 ◽

Author(s):

Sai Ba ◽

Guangpeng Gao ◽

Tianhu Li ◽

Hao Zhang

Keyword(s):

Design Strategy ◽

Living Cells ◽

Molecular Probes ◽

Chemical Bonds ◽

Dna Topoisomerase ◽

Dna Topoisomerases ◽

Catalytic Activities ◽

New Strategy ◽

Clinical Tools ◽

Catalytic Cycles

Abstract Most activity-based molecular probes are designed to target enzymes that catalyze the breaking of chemical bonds and the conversion of a unimolecular substrate into bimolecular products. However, DNA topoisomerases are a class of enzymes that alter DNA topology without producing any molecular segments during catalysis, which hinders the development of practical methods for diagnosing these key biomarkers in living cells. Here, we established a new strategy for the effective sensing of the expression levels and catalytic activities of topoisomerases in cell-free systems and human cells. Using our newly designed biosensors, we tricked DNA topoisomerases within their catalytic cycles to switch on fluorescence and resume new rounds of catalysis. Considering that human topoisomerases have been widely recognized as biomarkers for multiple cancers and identified as promising targets for several anticancer drugs, we believe that these DNA-based biosensors and our design strategy would greatly benefit the future development of clinical tools for cancer diagnosis and treatment.

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Fluorophores and Their Applications as Molecular Probes in Living Cells

Current Organic Chemistry ◽

10.2174/1385272811317060003 ◽

2013 ◽

Vol 17 (6) ◽

pp. 564-579 ◽

Cited By ~ 34

Author(s):

Junjia Liu ◽

Chen Liu ◽

Wei He

Keyword(s):

Living Cells ◽

Molecular Probes

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Interaction of molecular probes with living cells and tissues. Part 2

Histochemistry ◽

10.1007/bf00266632 ◽

1990 ◽

Vol 94 (3) ◽

Cited By ~ 43

Author(s):

F. Rashid ◽

R.W. Horobin

Keyword(s):

Living Cells ◽

Molecular Probes

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Oligonucleotides DNA containing 8-trifluoromethyl-2′-deoxyguanosine for observing Z-DNA structure

Nucleic Acids Research ◽

10.1093/nar/gkaa505 ◽

2020 ◽

Cited By ~ 1

Author(s):

Hong-Liang Bao ◽

Tatsuki Masuzawa ◽

Takanori Oyoshi ◽

Yan Xu

Keyword(s):

Genetic Diseases ◽

Dna Structure ◽

2D Nmr ◽

Living Cells ◽

Molecular Probes ◽

Alternative Form ◽

Left Handed ◽

Z Dna ◽

And Function

Abstract Z-DNA is known to be a left-handed alternative form of DNA and has important biological roles as well as being related to cancer and other genetic diseases. It is therefore important to investigate Z-DNA structure and related biological events in living cells. However, the development of molecular probes for the observation of Z-DNA structures inside living cells has not yet been realized. Here, we have succeeded in developing site-specific trifluoromethyl oligonucleotide DNA by incorporation of 8-trifluoromethyl-2′-deoxyguanosine (FG). 2D NMR strongly suggested that FG adopted a syn conformation. Trifluoromethyl oligonucleotides dramatically stabilized Z-DNA, even under physiological salt concentrations. Furthermore, the trifluoromethyl DNA can be used to directly observe Z-form DNA structure and interaction of DNA with proteins in vitro, as well as in living human cells by19F NMR spectroscopy for the first time. These results provide valuable information to allow understanding of the structure and function of Z-DNA.

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Poly(ADP-ribose) polymers regulate DNA topoisomerase I (Top1) nuclear dynamics and camptothecin sensitivity in living cells

Nucleic Acids Research ◽

10.1093/nar/gkw665 ◽

2016 ◽

Vol 44 (17) ◽

pp. 8363-8375 ◽

Cited By ~ 26

Author(s):

Subhendu K. Das ◽

Ishita Rehman ◽

Arijit Ghosh ◽

Souvik Sengupta ◽

Papiya Majumdar ◽

...

Keyword(s):

Topoisomerase I ◽

Living Cells ◽

Dna Topoisomerase I ◽

Nuclear Dynamics ◽

Dna Topoisomerase

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The SARS CoV-2 spike domain, RGD and integrin binding effect-relationship for vaccine design strategy

Annals of Proteomics and Bioinformatics ◽

10.29328/journal.apb.1001014 ◽

2021 ◽

Vol 5 (1) ◽

pp. 027-041

Author(s):

M Luisetto ◽

G Tarro ◽

Edbey Khaled ◽

N Almukthar ◽

L Cabianca ◽

...

Keyword(s):

Vaccine Design ◽

Design Strategy ◽

Spike Protein ◽

Pathological Effect ◽

Technical Data ◽

Binding Effect ◽

New Strategy ◽

Rare Side Effect ◽

New Generation

Related the need to search new strategy in vaccine design in order to reduce also some rare effect like trombosys for some registered products it is interesting the role played by the SPIKE RGD domain. The binding with molecules like Fibronectin is a process that must to be deeply investigated. A better understanding in this process can be used to improve safety of the new generation of COVID vaccine. The rare effect like thrombosis recognized by regulatory agency produced a modification of technical data sheet of some vaccine so the phenomena Is interesting to be more investigated. Spike protein and its domains are involved in producing pathological effect of the COVID-19 disease. What it is interesting is that some pathological effect of this pathology are similar to some rare side effect produced by some COVID-19 vaccine classes. After a review of interesting literature related this topics is submitted an experimental projects able to verify in vitro the spike procoaugulant property.

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Cr-Phthalocyanine Porous Organic Polymer as an Efficient and Selective Catalyst for Mono Carbonylation of Epoxides to Lactones

Catalysts ◽

10.3390/catal10080905 ◽

2020 ◽

Vol 10 (8) ◽

pp. 905

Author(s):

Vinothkumar Ganesan ◽

Sungho Yoon

Keyword(s):

Lewis Acid ◽

Design Strategy ◽

Organic Polymer ◽

Acid Base ◽

One Pot ◽

Selective Catalyst ◽

Catalytic Activities ◽

Porous Organic Polymer ◽

Polymeric Catalyst ◽

Highly Porous

A facile, one-pot design strategy to construct chromium(III)-phthalocyanine chlorides (Pc’CrCl) to form porous organic polymer (POP-Pc’CrCl) using solvent knitting Friedel-Crafts reaction (FCR) is described. The generated highly porous POP-Pc’CrCl is functionalized by post-synthetic exchange reaction with nucleophilic cobaltate ions to provide an heterogenized carbonylation catalyst (POP-Pc’CrCo(CO)4) with Lewis acid-base type bimetallic units. The produced porous polymeric catalyst is identical to that homogeneous counterpart in structure and coordination environments. The catalyst is very selective and effective for mono carbonylation of epoxide into corresponding lactone and the activities are comparable to those observed for a homogeneous Pc’CrCo(CO)4 catalyst. The (POP-Pc’CrCo(CO)4) also displayed a good catalytic activities and recyclability upon successive recycles.

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