Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>)
Here, we report a biochemical characterization of recombinant maize indole-3-acetyl-β-<span style="font-variant: small-caps;">d</span>-glucose (IAGlc) synthase which glucosylates indole-3-acetic acid (IAA) and thus abolishes its auxinic activity affecting plant hormonal homeost...
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MDPI AG
2021-03-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/22/7/3355 |
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| author | Anna Ciarkowska Maciej Ostrowski Anna Kozakiewicz |
| author_facet | Anna Ciarkowska Maciej Ostrowski Anna Kozakiewicz |
| author_sort | Anna Ciarkowska |
| collection | DOAJ |
| description | Here, we report a biochemical characterization of recombinant maize indole-3-acetyl-β-<span style="font-variant: small-caps;">d</span>-glucose (IAGlc) synthase which glucosylates indole-3-acetic acid (IAA) and thus abolishes its auxinic activity affecting plant hormonal homeostasis. Substrate specificity analysis revealed that IAA is a preferred substrate of IAGlc synthase; however, the enzyme can also glucosylate indole-3-butyric acid and indole-3-propionic acid with the relative activity of 66% and 49.7%, respectively. <i>K<sub>M</sub></i> values determined for IAA and UDP glucose are 0.8 and 0.7 mM, respectively. 2,4-Dichlorophenoxyacetic acid is a competitive inhibitor of the synthase and causes a 1.5-fold decrease in the enzyme affinity towards IAA, with the <i>K<sub>i</sub></i> value determined as 117 μM, while IAA–Asp acts as an activator of the synthase. Two sugar-phosphate compounds, ATP and glucose-1-phosphate, have a unique effect on the enzyme by acting as activators at low concentrations and showing inhibitory effect at higher concentrations (above 0.6 and 4 mM for ATP and glucose-1-phosphate, respectively). Results of molecular docking revealed that both compounds can bind to the PSPG (plant secondary product glycosyltransferase) motif of IAGlc synthase; however, there are also different potential binding sites present in the enzyme. We postulate that IAGlc synthase may contain more than one binding site for ATP and glucose-1-phosphate as reflected in its activity modulation. |
| first_indexed | 2024-03-10T12:55:29Z |
| format | Article |
| id | doaj.art-c312b30689e644d8bda681d964794b7c |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T12:55:29Z |
| publishDate | 2021-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-c312b30689e644d8bda681d964794b7c2023-11-21T11:57:34ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-03-01227335510.3390/ijms22073355Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>)Anna Ciarkowska0Maciej Ostrowski1Anna Kozakiewicz2Faculty of Biological and Veterinary Sciences, Nicolaus Copernicus University in Toruń, 87-100 Toruń, PolandFaculty of Biological and Veterinary Sciences, Nicolaus Copernicus University in Toruń, 87-100 Toruń, PolandFaculty of Chemistry, Nicolaus Copernicus University in Toruń, 87-100 Toruń, PolandHere, we report a biochemical characterization of recombinant maize indole-3-acetyl-β-<span style="font-variant: small-caps;">d</span>-glucose (IAGlc) synthase which glucosylates indole-3-acetic acid (IAA) and thus abolishes its auxinic activity affecting plant hormonal homeostasis. Substrate specificity analysis revealed that IAA is a preferred substrate of IAGlc synthase; however, the enzyme can also glucosylate indole-3-butyric acid and indole-3-propionic acid with the relative activity of 66% and 49.7%, respectively. <i>K<sub>M</sub></i> values determined for IAA and UDP glucose are 0.8 and 0.7 mM, respectively. 2,4-Dichlorophenoxyacetic acid is a competitive inhibitor of the synthase and causes a 1.5-fold decrease in the enzyme affinity towards IAA, with the <i>K<sub>i</sub></i> value determined as 117 μM, while IAA–Asp acts as an activator of the synthase. Two sugar-phosphate compounds, ATP and glucose-1-phosphate, have a unique effect on the enzyme by acting as activators at low concentrations and showing inhibitory effect at higher concentrations (above 0.6 and 4 mM for ATP and glucose-1-phosphate, respectively). Results of molecular docking revealed that both compounds can bind to the PSPG (plant secondary product glycosyltransferase) motif of IAGlc synthase; however, there are also different potential binding sites present in the enzyme. We postulate that IAGlc synthase may contain more than one binding site for ATP and glucose-1-phosphate as reflected in its activity modulation.https://www.mdpi.com/1422-0067/22/7/3355auxin conjugateindole-3-acetic acidUDPG-dependent IAA glucosyltransferaseenzyme modulatorsinhibitor–enzyme interaction |
| spellingShingle | Anna Ciarkowska Maciej Ostrowski Anna Kozakiewicz Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>) International Journal of Molecular Sciences auxin conjugate indole-3-acetic acid UDPG-dependent IAA glucosyltransferase enzyme modulators inhibitor–enzyme interaction |
| title | Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>) |
| title_full | Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>) |
| title_fullStr | Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>) |
| title_full_unstemmed | Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>) |
| title_short | Biochemical Characterization of Recombinant UDPG-Dependent IAA Glucosyltransferase from Maize (<i>Zea mays</i>) |
| title_sort | biochemical characterization of recombinant udpg dependent iaa glucosyltransferase from maize i zea mays i |
| topic | auxin conjugate indole-3-acetic acid UDPG-dependent IAA glucosyltransferase enzyme modulators inhibitor–enzyme interaction |
| url | https://www.mdpi.com/1422-0067/22/7/3355 |
| work_keys_str_mv | AT annaciarkowska biochemicalcharacterizationofrecombinantudpgdependentiaaglucosyltransferasefrommaizeizeamaysi AT maciejostrowski biochemicalcharacterizationofrecombinantudpgdependentiaaglucosyltransferasefrommaizeizeamaysi AT annakozakiewicz biochemicalcharacterizationofrecombinantudpgdependentiaaglucosyltransferasefrommaizeizeamaysi |