CEFOBI   05405
CENTRO DE ESTUDIOS FOTOSINTETICOS Y BIOQUIMICOS
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Structural and Biophysical Characterization of Phosphotransacetylase from
Autor/es:
VALERIA A. CAMPOS-BERMUDEZ, FEDERICO P. BOLOGNA , JAVIER M. GONZÁLEZ , EDWIN POZHARSKI , MARÍA FABIANA DRINCOVICH AND CARLOS S. ANDREO.
Lugar:
Salta-Argentina
Reunión:
Congreso; Latin American Protein Society Meeting; 2010
Resumen:
Escherichia coli phosphotransacetylase (Pta) catalyses the reversible inter-conversion of acetyl-CoA and acetyl-P. Both compounds are critical in E. coli metabolism, whereas acetyl-P is also involved in the regulation of certain signal transduction pathways. Along with acetate kinase, Pta plays an important role in acetate production when E. coli grows on rich medium; alternatively, it is involved in acetate utilization at high acetate concentration. E. coli Pta is composed by three different domains, but only the C-terminal one, called PTA_PTB, is specific for all Ptas and the unique domain in certain Ptas. The characterization of three E. coli Pta truncated proteins, which all display Pta activity, indicates that the substrate binding site is located at the C-terminal PTA_PTB domain. However, the N-terminal P-loop NTPase domain is involved in the expression of the maximal catalytic activity, in the stabilization of the hexameric native state and in Pta activity regulation by NADH, ATP, PEP and pyruvate. To better characterize the effectors and substrate binding sites on this enzyme a quenching fluorescence approach has been carried out. From this technique we are able to determine the dissociation constants for each ligand and its temperature dependence and the presence of other effectors. Additionally, crystals of Pta have been obtained in commercially available crystallization screens, which diffracted to ~ 7 Å. Crystal optimization trials are underway; which may allow us to solve the structure of Pta, currently unknown.