Asunto(s)
Envejecimiento , Células Madre Hematopoyéticas/química , Factores Reguladores del Interferón/análisis , ADP-Ribosil Ciclasa 1 , Adolescente , Adulto , Factores de Edad , Anciano , Animales , Antígenos CD34 , Humanos , Factores Reguladores del Interferón/genética , Ratones , Persona de Mediana Edad , Adulto JovenRESUMEN
Carbonic anhydrases fall into three distinct evolutionary and structural classes: alpha, beta, and gamma. The beta-class carbonic anhydrases (beta-CAs) are widely distributed among higher plants, simple eukaryotes, eubacteria, and archaea. We have determined the crystal structure of ECCA, a beta-CA from Escherichia coli, to a resolution of 2.0 A. In agreement with the structure of the beta-CA from the chloroplast of the red alga Porphyridium purpureum, the active-site zinc in ECCA is tetrahedrally coordinated by the side chains of four conserved residues. These results confirm the observation of a unique pattern of zinc ligation in at least some beta-CAS: The absence of a water molecule in the inner coordination sphere is inconsistent with known mechanisms of CA activity. ECCA activity is highly pH-dependent in the physiological range, and its expression in yeast complements an oxygen-sensitive phenotype displayed by a beta-CA-deletion strain. The structural and biochemical characterizations of ECCA presented here and the comparisons with other beta-CA structures suggest that ECCA can adopt two distinct conformations displaying widely divergent catalytic rates.
Asunto(s)
Anhidrasas Carbónicas/química , Escherichia coli/enzimología , Secuencia de Aminoácidos , Sitios de Unión , Anhidrasas Carbónicas/aislamiento & purificación , Anhidrasas Carbónicas/metabolismo , Cristalografía por Rayos X , Eliminación de Gen , Prueba de Complementación Genética , Concentración de Iones de Hidrógeno , Modelos Moleculares , Datos de Secuencia Molecular , Pliegue de Proteína , Estructura Cuaternaria de Proteína , Estructura Secundaria de Proteína , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Relación Estructura-Actividad , Zinc/metabolismoRESUMEN
Carbonic anhydrases are zinc metalloenzymes that fall into three distinct evolutionary and structural classes, alpha, beta and gamma. Although alpha-class enzymes, particularly mammalian carbonic anhydrase II, have been the subject of extensive structural studies, for the beta class, consisting of a wide variety of prokaryotic and plant chloroplast carbonic anhydrases, the structural data is quite limited. A member of the beta class from E. coli (CynT2) has been crystallized in native and selenomethionine-labelled forms and multiwavelength anomalous dispersion techniques have been applied in order to determine the positions of anomalous scatterers. The resulting phase information is sufficient to produce an interpretable electron-density map. A crystal structure for CynT2 would contribute significantly to the emerging structural knowledge of a biologically important class of enzymes that perform critical functions in carbon fixation and prokaryotic metabolism.
