| Structural analysis of K(+) dependence in L: -asparaginases from Lotus japonicus. | |
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MedLine Citation:
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PMID: 21390508 Owner: NLM Status: Publisher |
Abstract/OtherAbstract:
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The molecular features responsible for the existence in plants of K(+)-dependent asparaginases have been investigated. For this purpose, two different cDNAs were isolated in Lotus japonicus, encoding for K(+)-dependent (LjNSE1) or K(+)-independent (LjNSE2) asparaginases. Recombinant proteins encoded by these cDNAs have been purified and characterized. Both types of asparaginases are composed by two different subunits, α (20 kDa) and β (17 kDa), disposed as (αβ)(2) quaternary structure. Major differences were found in the catalytic efficiency of both enzymes, due to the fact that K(+) is able to increase by tenfold the enzyme activity and lowers the K (m) for asparagine specifically in LjNSE1 but not in LjNSE2 isoform. Optimum LjNSE1 activity was found at 5-50 mM K(+), with a K (m) for K(+) of 0.25 mM. Na(+) and Rb(+) can, to some extent, substitute for K(+) on the activating effect of LjNSE1 more efficiently than Cs(+) and Li(+) does. In addition, K(+) is able to stabilize LjNSE1 against thermal inactivation. Protein homology modelling and molecular dynamics studies, complemented with site-directed mutagenesis, revealed the key importance of E248, D285 and E286 residues for the catalytic activity and K(+) dependence of LjNSE1, as well as the crucial relevance of K(+) for the proper orientation of asparagine substrate within the enzyme molecule. On the other hand, LjNSE2 but not LjNSE1 showed β-aspartyl-hydrolase activity (K (m) = 0.54 mM for β-Asp-His). These results are discussed in terms of the different physiological significance of these isoenzymes in plants. |
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Authors:
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Alfredo Credali; Antonio Díaz-Quintana; Margarita García-Calderón; Miguel A De la Rosa; Antonio J Márquez; José M Vega |
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Publication Detail:
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Type: JOURNAL ARTICLE Date: 2011-3-10 |
Journal Detail:
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Title: Planta Volume: - ISSN: 1432-2048 ISO Abbreviation: - Publication Date: 2011 Mar |
Date Detail:
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Created Date: 2011-3-10 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 1250576 Medline TA: Planta Country: - |
Other Details:
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Languages: ENG Pagination: - Citation Subset: - |
Affiliation:
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Departamento de Bioquímica Vegetal y Biología Molecular, Facultad de Química, Universidad de Sevilla, c/Prof. García González, nº 1, Apartado 1203, 41071, Seville, Spain. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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