Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i>
The green unicellular algae <i>Chlamydomonas reinhardtii</i> contains 12–13 carbonic anhydrases (CAs). For a long time, the two closely related α-CAs of the periplasmic membrane CAH1 and CAH2 were considered to be the CAs with the highest CO<sub>2</sub> hydration activity. Th...
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2024-12-01
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| author | Vasily V. Terentyev Liubov I. Trubitsina Anna K. Shukshina Ivan V. Trubitsin Natalia N. Rudenko |
| author_facet | Vasily V. Terentyev Liubov I. Trubitsina Anna K. Shukshina Ivan V. Trubitsin Natalia N. Rudenko |
| author_sort | Vasily V. Terentyev |
| collection | DOAJ |
| description | The green unicellular algae <i>Chlamydomonas reinhardtii</i> contains 12–13 carbonic anhydrases (CAs). For a long time, the two closely related α-CAs of the periplasmic membrane CAH1 and CAH2 were considered to be the CAs with the highest CO<sub>2</sub> hydration activity. The recombinant protein α-CA CAH3 (rCAH3) from the thylakoid lumen obtained in the present study showed more than three times higher activity compared to CAH1 and more than 11 times higher compared to previous studies with rCAH3. Long-term sustainability of the enzyme was observed at alkaline pH (>8), with maintenance of half of its activity at 4 °C for up to 50 days. Thermostability of rCAH3 indicated the retention of the activity at 20 °C for one hour at pH 9–10 with its ~50% decrease at pH 6–7. However, the residual activity of rCAH3 after incubation at an extremely high temperature (75 °C) for 15 min led to the formation of the double-hump graph with maxima at pH 6 and 9. The enzyme demonstrated high sensitivity to ethoxyzolamide and acetazolamide at nM concentrations, to Zn<sup>2+</sup> and Cu<sup>2+</sup> cations at 1 mM concentrations, and L-cysteine was able to completely inhibit CA activity of rCAH3 through reduction of sulfhydryl groups. Esterase activity of rCAH3 was well detected with values comparable to those of bovine CAII, but with a maximum at pH 8 instead of pH 9, which is usual for bovine CAII. The results indicated that CAH3 may be the most active CA of <i>C. reinhardtii</i> and that its role in the photosynthetic apparatus function could have been underestimated in previous works. |
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| institution | Kabale University |
| issn | 2223-7747 |
| language | English |
| publishDate | 2024-12-01 |
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| series | Plants |
| spelling | doaj-art-f870e59b49394411907909416988ad882025-01-10T13:19:36ZengMDPI AGPlants2223-77472024-12-011415510.3390/plants14010055Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i>Vasily V. Terentyev0Liubov I. Trubitsina1Anna K. Shukshina2Ivan V. Trubitsin3Natalia N. Rudenko4Institute of Basic Biological Problems, Federal Research Center “Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences”, 142290 Pushchino, RussiaG.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms, Federal Research Center “Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences”, 142290 Pushchino, RussiaInstitute of Basic Biological Problems, Federal Research Center “Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences”, 142290 Pushchino, RussiaG.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms, Federal Research Center “Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences”, 142290 Pushchino, RussiaInstitute of Basic Biological Problems, Federal Research Center “Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences”, 142290 Pushchino, RussiaThe green unicellular algae <i>Chlamydomonas reinhardtii</i> contains 12–13 carbonic anhydrases (CAs). For a long time, the two closely related α-CAs of the periplasmic membrane CAH1 and CAH2 were considered to be the CAs with the highest CO<sub>2</sub> hydration activity. The recombinant protein α-CA CAH3 (rCAH3) from the thylakoid lumen obtained in the present study showed more than three times higher activity compared to CAH1 and more than 11 times higher compared to previous studies with rCAH3. Long-term sustainability of the enzyme was observed at alkaline pH (>8), with maintenance of half of its activity at 4 °C for up to 50 days. Thermostability of rCAH3 indicated the retention of the activity at 20 °C for one hour at pH 9–10 with its ~50% decrease at pH 6–7. However, the residual activity of rCAH3 after incubation at an extremely high temperature (75 °C) for 15 min led to the formation of the double-hump graph with maxima at pH 6 and 9. The enzyme demonstrated high sensitivity to ethoxyzolamide and acetazolamide at nM concentrations, to Zn<sup>2+</sup> and Cu<sup>2+</sup> cations at 1 mM concentrations, and L-cysteine was able to completely inhibit CA activity of rCAH3 through reduction of sulfhydryl groups. Esterase activity of rCAH3 was well detected with values comparable to those of bovine CAII, but with a maximum at pH 8 instead of pH 9, which is usual for bovine CAII. The results indicated that CAH3 may be the most active CA of <i>C. reinhardtii</i> and that its role in the photosynthetic apparatus function could have been underestimated in previous works.https://www.mdpi.com/2223-7747/14/1/55<i>Chlamydomonas</i>carbonic anhydraseCAH3photosystem IIrecombinant protein |
| spellingShingle | Vasily V. Terentyev Liubov I. Trubitsina Anna K. Shukshina Ivan V. Trubitsin Natalia N. Rudenko Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i> Plants <i>Chlamydomonas</i> carbonic anhydrase CAH3 photosystem II recombinant protein |
| title | Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i> |
| title_full | Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i> |
| title_fullStr | Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i> |
| title_full_unstemmed | Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i> |
| title_short | Highly Active Carbonic Anhydrase of the Thylakoid Lumen of <i>Chlamydomonas reinhardtii</i> |
| title_sort | highly active carbonic anhydrase of the thylakoid lumen of i chlamydomonas reinhardtii i |
| topic | <i>Chlamydomonas</i> carbonic anhydrase CAH3 photosystem II recombinant protein |
| url | https://www.mdpi.com/2223-7747/14/1/55 |
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