Kinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidases
Enzymatic O2 sensors transduce the availability of O2 within the cell into a physiological, typically adaptive response. One such O2-sensing enzymatic family is the N-terminal cysteine dioxygenases in plants (plant cysteine oxidases [PCOs]). In vitro kinetic studies have determined the O2-sensing ca...
| Main Authors: | , , |
|---|---|
| Other Authors: | |
| Format: | Book section |
| Language: | English |
| Published: |
Springer Nature
2023
|
| _version_ | 1826312733499326464 |
|---|---|
| author | Dirr, A Gunawardana, DM Flashman, E |
| author2 | Weinert, EE |
| author_facet | Weinert, EE Dirr, A Gunawardana, DM Flashman, E |
| author_sort | Dirr, A |
| collection | OXFORD |
| description | Enzymatic O2 sensors transduce the availability of O2 within the cell into a physiological, typically adaptive response. One such O2-sensing enzymatic family is the N-terminal cysteine dioxygenases in plants (plant cysteine oxidases [PCOs]). In vitro kinetic studies have determined the O2-sensing capacity of PCOs. Here we describe the rationale and experimental protocol for an assay with which the O2 sensitivity of Arabidopsis thaliana PCOs (AtPCOs) can be measured. We explain each step from the recombinant protein synthesis of AtPCOs to the steady-state kinetic assays of AtPCOs for primary substrate and O2 from which kinetic parameters can be derived. The same techniques can be applied to other N-terminal cysteine thiol dioxygenases, e.g. 2-aminoethanethiol dioxygenase (ADO), and similar principles can be applied to determine kinetic characteristics of other oxygenase enzymes towards O2. |
| first_indexed | 2024-03-07T08:26:28Z |
| format | Book section |
| id | oxford-uuid:97fa735f-26b5-45cd-9f75-8857f0586737 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-04-23T08:26:03Z |
| publishDate | 2023 |
| publisher | Springer Nature |
| record_format | dspace |
| spelling | oxford-uuid:97fa735f-26b5-45cd-9f75-8857f05867372024-04-12T09:36:23ZKinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidasesBook sectionhttp://purl.org/coar/resource_type/c_1843uuid:97fa735f-26b5-45cd-9f75-8857f0586737EnglishSymplectic ElementsSpringer Nature2023Dirr, AGunawardana, DMFlashman, EWeinert, EEEnzymatic O2 sensors transduce the availability of O2 within the cell into a physiological, typically adaptive response. One such O2-sensing enzymatic family is the N-terminal cysteine dioxygenases in plants (plant cysteine oxidases [PCOs]). In vitro kinetic studies have determined the O2-sensing capacity of PCOs. Here we describe the rationale and experimental protocol for an assay with which the O2 sensitivity of Arabidopsis thaliana PCOs (AtPCOs) can be measured. We explain each step from the recombinant protein synthesis of AtPCOs to the steady-state kinetic assays of AtPCOs for primary substrate and O2 from which kinetic parameters can be derived. The same techniques can be applied to other N-terminal cysteine thiol dioxygenases, e.g. 2-aminoethanethiol dioxygenase (ADO), and similar principles can be applied to determine kinetic characteristics of other oxygenase enzymes towards O2. |
| spellingShingle | Dirr, A Gunawardana, DM Flashman, E Kinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidases |
| title | Kinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidases |
| title_full | Kinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidases |
| title_fullStr | Kinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidases |
| title_full_unstemmed | Kinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidases |
| title_short | Kinetic measurements to investigate the oxygen-sensing properties of plant cysteine oxidases |
| title_sort | kinetic measurements to investigate the oxygen sensing properties of plant cysteine oxidases |
| work_keys_str_mv | AT dirra kineticmeasurementstoinvestigatetheoxygensensingpropertiesofplantcysteineoxidases AT gunawardanadm kineticmeasurementstoinvestigatetheoxygensensingpropertiesofplantcysteineoxidases AT flashmane kineticmeasurementstoinvestigatetheoxygensensingpropertiesofplantcysteineoxidases |