Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD protein
Azospirillum brasilense is a diazotroph found in association with important agricultural crops. In this organism, the regulation of nitrogen fixation by ammonium ions involves several proteins including the uridylyltransferase/uridylyl-removing enzyme, GlnD, which reversibly uridylylates the two PII...
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| Format: | Article |
| Language: | English |
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Associação Brasileira de Divulgação Científica
2008-04-01
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| Series: | Brazilian Journal of Medical and Biological Research |
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| Online Access: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2008000400006 |
| _version_ | 1818032783995437056 |
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| author | L.M. Araújo L.F. Huergo A.L. Invitti C.I. Gimenes A.C. Bonatto R.A. Monteiro E.M. Souza F.O. Pedrosa L.S. Chubatsu |
| author_facet | L.M. Araújo L.F. Huergo A.L. Invitti C.I. Gimenes A.C. Bonatto R.A. Monteiro E.M. Souza F.O. Pedrosa L.S. Chubatsu |
| author_sort | L.M. Araújo |
| collection | DOAJ |
| description | Azospirillum brasilense is a diazotroph found in association with important agricultural crops. In this organism, the regulation of nitrogen fixation by ammonium ions involves several proteins including the uridylyltransferase/uridylyl-removing enzyme, GlnD, which reversibly uridylylates the two PII proteins, GlnB and GlnZ, in response to the concentration of ammonium ions. In the present study, the uridylylation/deuridylylation cycle of A. brasilense GlnB and GlnZ proteins by GlnD was reconstituted in vitro using the purified proteins. The uridylylation assay was analyzed using non-denaturing polyacrylamide gel electrophoresis and fluorescent protein detection. Our results show that the purified A. brasilense GlnB and GlnZ proteins were uridylylated by the purified A. brasilense GlnD protein in a process dependent on ATP and 2-oxoglutarate. The dependence on ATP for uridylylation was similar for both proteins. On the other hand, at micromolar concentration of 2-oxoglutarate (up to 100 µM), GlnB uridylylation was almost twice that of GlnZ, an effect that was not observed at higher concentrations of 2-oxoglutarate (up to 10 mM). Glutamine inhibited uridylylation and stimulated deuridylylation of both GlnB and GlnZ. However, glutamine seemed to inhibit GlnZ uridylylation more efficiently. Our results suggest that the differences in the uridylylation pattern of GlnB and GlnZ might be important for fine-tuning of the signaling pathway of cellular nitrogen status in A. brasilense. |
| first_indexed | 2024-12-10T06:12:52Z |
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| institution | Directory Open Access Journal |
| issn | 0100-879X 1414-431X |
| language | English |
| last_indexed | 2024-12-10T06:12:52Z |
| publishDate | 2008-04-01 |
| publisher | Associação Brasileira de Divulgação Científica |
| record_format | Article |
| series | Brazilian Journal of Medical and Biological Research |
| spelling | doaj.art-d979220892b74f1cb862d8aa5e42f5be2022-12-22T01:59:31ZengAssociação Brasileira de Divulgação CientíficaBrazilian Journal of Medical and Biological Research0100-879X1414-431X2008-04-01414289294Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD proteinL.M. AraújoL.F. HuergoA.L. InvittiC.I. GimenesA.C. BonattoR.A. MonteiroE.M. SouzaF.O. PedrosaL.S. ChubatsuAzospirillum brasilense is a diazotroph found in association with important agricultural crops. In this organism, the regulation of nitrogen fixation by ammonium ions involves several proteins including the uridylyltransferase/uridylyl-removing enzyme, GlnD, which reversibly uridylylates the two PII proteins, GlnB and GlnZ, in response to the concentration of ammonium ions. In the present study, the uridylylation/deuridylylation cycle of A. brasilense GlnB and GlnZ proteins by GlnD was reconstituted in vitro using the purified proteins. The uridylylation assay was analyzed using non-denaturing polyacrylamide gel electrophoresis and fluorescent protein detection. Our results show that the purified A. brasilense GlnB and GlnZ proteins were uridylylated by the purified A. brasilense GlnD protein in a process dependent on ATP and 2-oxoglutarate. The dependence on ATP for uridylylation was similar for both proteins. On the other hand, at micromolar concentration of 2-oxoglutarate (up to 100 µM), GlnB uridylylation was almost twice that of GlnZ, an effect that was not observed at higher concentrations of 2-oxoglutarate (up to 10 mM). Glutamine inhibited uridylylation and stimulated deuridylylation of both GlnB and GlnZ. However, glutamine seemed to inhibit GlnZ uridylylation more efficiently. Our results suggest that the differences in the uridylylation pattern of GlnB and GlnZ might be important for fine-tuning of the signaling pathway of cellular nitrogen status in A. brasilense.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2008000400006Azospirillum brasilenseNitrogen fixationPII-like proteinGlnDGlnBGlnZ |
| spellingShingle | L.M. Araújo L.F. Huergo A.L. Invitti C.I. Gimenes A.C. Bonatto R.A. Monteiro E.M. Souza F.O. Pedrosa L.S. Chubatsu Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD protein Brazilian Journal of Medical and Biological Research Azospirillum brasilense Nitrogen fixation PII-like protein GlnD GlnB GlnZ |
| title | Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD protein |
| title_full | Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD protein |
| title_fullStr | Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD protein |
| title_full_unstemmed | Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD protein |
| title_short | Different responses of the GlnB and GlnZ proteins upon in vitro uridylylation by the Azospirillum brasilense GlnD protein |
| title_sort | different responses of the glnb and glnz proteins upon in vitro uridylylation by the azospirillum brasilense glnd protein |
| topic | Azospirillum brasilense Nitrogen fixation PII-like protein GlnD GlnB GlnZ |
| url | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2008000400006 |
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