Oxygen Sensing. Methods and Protocols.

0580389 - MBÚ 2024 RIV US eng M - Část monografie knihy
Vávra, J. - Sergunin, A. - Stráňava, M. - Kádek, Alan - Shimitzu, T. - Man, Petr - Martínková, M.
Hydrogen/Deuterium Exchange Mass Spectrometry of Heme-Based Oxygen Sensor Proteins.
Oxygen Sensing. Methods and Protocols.. New York: Humana, 2023 - (Weinert, E.), s. 99-122. ISBN 978-1-0716-3079-2
Grant CEP: GA MŠMT(CZ) ED1.1.00/02.0109
Výzkumná infrastruktura: CIISB II - 90127
Institucionální podpora: RVO:61388971
Klíčová slova: Hydrogen/deuterium exchange * Mass spectrometry * Protein conformational dynamics * Ligand binding * Signal transduction * Globin-coupled histidine kinase * Heme-containing oxygen sensors
Obor OECD: Biochemical research methods
https://link.springer.com/protocol/10.1007/978-1-0716-3080-8_8

Hydrogen/deuterium exchange (HDX) is a well-established analytical technique that enables monitoring of protein dynamics and interactions by probing the isotope exchange of backbone amides. It has virtually no limitations in terms of protein size, flexibility, or reaction conditions and can thus be performed in solution at different pH values and temperatures under controlled redox conditions. Thanks to its coupling with mass spectrometry (MS), it is also straightforward to perform and has relatively high throughput, making it an excellent complement to the high-resolution methods of structural biology. Given the recent expansion of artificial intelligence-aided protein structure modeling, there is considerable demand for techniques allowing fast and unambiguous validation of in silico predictions HDX-MS is well-placed to meet this demand. Here we present a protocol for HDX-MS and illustrate its use in characterizing the dynamics and structural changes of a dimeric heme-containing oxygen sensor protein as it responds to changes in its coordination and redox state. This allowed us to propose a mechanism by which the signal (oxygen binding to the heme iron in the sensing domain) is transduced to the protein’s functional domain.
Trvalý link: https://hdl.handle.net/11104/0349167