Thymidylate kinase (TMK) of the photosynthetic, nitrogen-fixing cyanobacterium Nostoc sp. strain PCC7120 : Biophysical, biochemical and physiological characterisation
Copyright © 2021 Elsevier Masson SAS. All rights reserved.
| Veröffentlicht in: | Plant physiology and biochemistry : PPB. - 1991. - 166(2021) vom: 01. Sept., Seite 416-426 |
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| Format: | Online-Aufsatz |
| Sprache: | English |
| Veröffentlicht: |
2021
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| Zugriff auf das übergeordnete Werk: | Plant physiology and biochemistry : PPB |
| Schlagworte: | Journal Article Cyanobacteria Isothermal titration calorimetry Luciferase-based TMK assay Nostoc PCC7120 Photosystem II Thymidylate kinase Nucleoside-Phosphate Kinase EC 2.7.4.4 dTMP kinase mehr... |
| Zusammenfassung: | Copyright © 2021 Elsevier Masson SAS. All rights reserved. Thymidylate kinase (TMK/TMPK) is an important enzyme in DNA biosynthesis and catalyses the conversion of dTMP to dTDP. Due to its therapeutic potential, the focus has been on characterizing the TMK proteins of pathogens and human origin, with very little information available on the TMK proteins of photosynthetic organisms and agriculturally important nitrogen-fixing organisms. In this work we report the characterisation of TMK in an evolutionarily ancient organism, cyanobacteria. The TMK protein of the photosynthetic, nitrogen-fixing cyanobacterium Nostoc PCC7120 (AnTMK) was found to have low conformational stability, which related to its low Tm of ~46 °C confirmed by Differential Scanning Fluorimetry (DSF) and Differential Scanning Calorimetry (DSC) techniques. The AnTMK protein exhibited substrate specificity for dTMP and ATP with Km of 20.74 ± 1.47 μM and 20.17 ± 2.96 μM respectively. The enzyme kinetics data and the positive co-operativity observed between dTMP and ATP binding correlated well with the data obtained from Isothermal Titration Calorimetry (ITC). Homology model of the enzyme suggested that the binding mode of substrate nucleotides to the enzyme is conserved. When overexpressed constitutively in Nostoc PCC7120 (Antmk+), it supported faster growth measured in terms of chlorophyll a content under normal growth conditions, but exhibited lower photosynthetic efficiency. Compared to the vector control recombinant Nostoc AnpAM, the Antmk + cells exhibited higher photoinhibition at higher light irradiance with more open reaction centres and lower dissipation of heat, indicative of damage to photosynthetic machinery. This indicated that the TMK is likely to have a significant role in photosynthetic organisms |
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| Beschreibung: | Date Completed 07.09.2021 Date Revised 07.09.2021 published: Print-Electronic Citation Status MEDLINE |
| ISSN: | 1873-2690 |
| DOI: | 10.1016/j.plaphy.2021.06.019 |