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Tytuł:
Visualization of proteolytic enzymes using mass cytometry : compatible chemical probes
Wizualizacja enzymów proteolitycznych za pomocą sond chemicznych oraz cytometrii masowej
Autorzy:
Groborz, Katarzyna
Poręba, Marcin
Powiązania:
https://bibliotekanauki.pl/articles/2057909.pdf
Data publikacji:
2021
Wydawca:
Polskie Towarzystwo Chemiczne
Tematy:
sonda chemiczna
cytometria masowa
enzymy proteolityczne
nowotwory
śmierć komórki
chemical probe
mass cytometry
proteolytic enzymes
cancer
cell death
Opis:
Proteolytic enzymes, also known as peptidases or proteases, are protein catalysts that are primarily responsible for the hydrolysis of a peptide (amide) bond in peptide and protein substrates. By selective hydrolysis of selected substrates, these enzymes control many physiologically important processes including programmed cell death, blood coagulation cascade, protein maturation, fibrinolysis and many others. On the other hand, however, the imbalance in proteases activity leads to the development of diseases, including cancer, neurodegenerative diseases and coronary diseases etc.. In recent decades there has been great progress in studying the biological functions of many proteolytic enzymes. These observations were made possible through the use of various research techniques including genomics, epigenomics and proteomics. However, a major limitation of these techniques is the lack of information about the exact catalytic activity of the enzymes. For this reason, chemical probes are the most convenient toll for functional investigation of proteolytic enzymes. According to the generally accepted convention, chemical probes are compounds that can detect the catalytic activity of proteolytic enzymes. In general, chemical-based probes (activity-based probes, ABPs) consist of three main components: (1) a reactive binding group that binds permanently to the enzyme active site, (2) a recognition sequence (usually a peptide), which is responsible for the selective binding of a given probe to an individual enzyme or group of enzymes, and (3) a tag, mainly a fluorophore, enabling for detection of the probe-enzyme complex. However, the current limitation of ABPs is that only up to four enzymes can be detected and visualized in parallel, which significantly impedes their application for multi-parametric analysis. To date, the detection of proteases with the use of ABPs was limited to individual enzymes being investigated one by one, thus the obtained picture was far from being complete. In this review we describe the development of a new type of enzyme ABPs, so called TOF-probes that are compatible with mass cytometry format. The application of metal isotopes instead of fluorophores, makes possible to significantly increase the number of enzymes, which can be simultaneously visualized using chemical probes. Mass cytometry is a revolutionary technology that adopts atomic mass spectrometry into flow cytometry applications. The excellence of this method is that each metal isotope (mostly from lanthanides) has its own peak on mass spectrum, which eliminates the problem of signal overlap, thus allows for monitoring of more than 40 parameters at single cell level.
Źródło:
Wiadomości Chemiczne; 2021, 75, 11-12; 1171--1191
0043-5104
2300-0295
Pojawia się w:
Wiadomości Chemiczne
Dostawca treści:
Biblioteka Nauki
Artykuł
Tytuł:
Narzędzia chemiczne do badania enzymów proteolitycznych przy użyciu cytometrii masowej
Mass cytometry-compatible chemical probes for the investigation of proteolytic enzymes
Autorzy:
Groborz, Katarzyna
Powiązania:
https://bibliotekanauki.pl/articles/2200437.pdf
Data publikacji:
2022
Wydawca:
Polskie Towarzystwo Chemiczne
Tematy:
markery chemiczne
cytometria masowa
enzymy proteolityczne
nowotwór
śmierć komórki
activity-based probes
mass cytometry
proteolytic enzymes
cancer
cell death
Opis:
Mass cytometry is one of the newest and most high-throughput technologies that allows for the investigation of complex biological systems at single cell level. It relies on the use of stable metal isotopes as labels of specific cell markers and therefore, allows for simultaneous analysis of more than 40 parameters at single cell level. In order to fully explore the potential of mass cytometry, researchers are trying to develop new experimental setups based on the application of pure metal isotopes in biological studies. The incorporation of antibodies into mass cytometry setups, while extremely selective and well-validated, limits the analysis as it shows the whole protein pool present in the cell. In our group, we developed new technology that allows for the identification of active forms of proteins-the ones that actively participate in cell signaling pathways. Activity-based probes are the most valuable tools for enzyme activity profiling and for years now they have been in the center of the method called Activity-Based Protein Profiling. Classic activity-based probes consist of three parts: a warhead (electrophilic binding group that covalently modifies enzyme active site), linker (specific peptide sequence or non-specific carbon chain) and the fluorescent tag that allows for enzyme detection and localization inside the cell. Spectral properties of commercially available fluorophores allow for the detection of up to dozen different cell parameters, with the use of various techniques such as confocal microscopy or flow cytometry. To increase the number of analyzed parameters, we designed activity-based probes that possess DOTA chelating moiety that is able to trap one metal atom per one probe. The combination of mass cytometry with highly selective activity-based probes allowed for the development of new technology that grants the possibility of multiparametric analysis of complex biological samples such as blood or cancer tissue. The new type of activity-based probes (so-called TOF-probes) incorporate various inhibitor scaffolds designed with HyCoSuL technology (Hybrid Combinatorial Substrate Libraries). These compounds possess a variety of unnatural amino acids in their structures, which significantly increases their selectivity toward proteases of interest.
Źródło:
Wiadomości Chemiczne; 2022, 76, 11-12; 864-880
0043-5104
2300-0295
Pojawia się w:
Wiadomości Chemiczne
Dostawca treści:
Biblioteka Nauki
Artykuł
    Wyświetlanie 1-2 z 2

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