Biochemical and structural characterization of Phosphotransacetylase from E. coli

CAMPOS BERMUDEZ VA; VALERIA A. CAMPOS-BERMUDEZ, FEDERICO PABLO BOLOGNA, MARÍA FABIANA DRINCOVICH AND CARLOS SANTIAGO ANDREO

San Miguel de Tucumán, Tucumán

Congreso; XLV Reunion Anual - Sociedad Argentina de Investigaión en Bioquímica y Biología molecular; 2009

SAIB

The successful adaptation of Escherichia coli, and other organisms, to constant nutritional changes, which range from feast to famine, relies primarily on the organism?s ability to ?switch off? carbon flux to acetate excretion and to ?turn on? acetate utilization. Acetate enters central metabolism as Ac-CoA, either directly through the activity of Ac-CoA synthetase (ACS) or in a two-step pathway involving phosphotrans-acetylase (PTA) and acetate kinase (AK). Sequence analysis of microbial genomes reveals two classes of Pta enzymes. Class I enzymes (PtaI) are 350 residues in length, whereas class II enzymes (PtaII) are twice as long as PtaI (700 residues). PtaI enzymes share end-to-end homology with the C-terminal domain of PtaII enzymes; hence, it is inferred that the active site of PtaII enzymes is located within their C-terminal domain. However, this assignment generate the question of what the function of the N-terminal domain of PtaII might be. Previous results from in vivo genetic and in vitro studies suggest that the Nterminal domain of Pta from Salmonella is a sensor for NADH and pyruvate. We evaluate this hypothesis through the study of three deletion mutants of Pta gene from E. coli. Kinetic analyses and the behavior against different metabolites reveal the essentiality of this additional domain on Pta