CEFOBI   05405
CENTRO DE ESTUDIOS FOTOSINTETICOS Y BIOQUIMICOS
Unidad Ejecutora - UE
artículos
Título:
Identification of domains involved in the allosteric regulation of cytosolic Arabidopsis thaliana NADP-malic enzymes
Autor/es:
GERRARD WHEELER, M. C.; ARIAS, C. L.; MAURINO, V. G.; ANDREO, C. S.; DRINCOVICH, M. F.
Revista:
FEBS JOURNAL
Editorial:
WILEY-BLACKWELL PUBLISHING, INC
Referencias:
Año: 2009 p. 5665 - 5677
ISSN:
1742-464X
Resumen:
<!-- /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-parent:""; margin:0in; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman"; mso-ansi-language:ES-AR;} @page Section1 {size:595.3pt 841.9pt; margin:70.85pt 85.05pt 70.85pt 85.05pt; mso-header-margin:35.4pt; mso-footer-margin:35.4pt; mso-paper-source:0;} div.Section1 {page:Section1;} --> The Arabidopsis thaliana genome contains four genes encoding NADP-malic enzymes (NADP-ME1 to -4). Two isoenzymes, NADP-ME2 and -3, which are shown to be located in the cytosol, share a remarkably high degree of identity (90%). However, they display different expression patterns and show distinct kinetic properties, especially with regard to their regulation by effectors, both in the forward -malate oxidative decarboxylation- and the reverse -pyruvate reductive carboxylation- reactions. In order to identify domains in the primary structure that could be responsible for the regulatory differences, four chimeras between these isoenzymes were constructed and analysed. All chimeric versions exhibited the same native structures as the parental proteins. Analysis of the chimeras constructed indicated that the region from amino acid residue 303 to the C-terminal end of NADP-ME2 is critical for fumarate activation. On the other hand, the region flanked by the amino acid residues 303 and 500 of NADP-ME3 is involved in the pH dependent-inhibition by high malate concentration. Furthermore, the N-terminal region of NADP-ME2 is necessary for the activation by succinate of the reverse reaction. Overall, the results show that NADP-ME2 and -3 are able to distinguish and interact differentially with similar C4 acids due to minimal structural differences. Therefore, although the active sites of NADP-ME2 and -3 are highly conserved, both isoenzymes acquired different allosteric sites, leading to the creation of proteins with unique regulatory mechanisms, probably best suited to the specific organ and developmental pattern of expression of each isoenzyme.