Characterization of aspartate aminotransferase from the cyanobacterium phormidiu lapideum

Abstract

Aspartate aminotransferase (AspAT) was purified to homogeneity from cell extracts of the non-N2-fixing cyanobacterium Phormidium lapideum. The NH2-terminal sequence of 25 amino acid residues was different from the sequences of the subfamily Iα of AspATs from eukaryotes and Escherichia coli, but it was similar to sequences of the subfamily Iγ of AspATs from archaebacteria and eubacteria. The enzyme was most active at 80°C and was stable at up to 75°C. Thermal inactivation (60-85°C) of the enzyme followed first-order kinetics, with 2-oxoglutarate causing a shift of the thermal inactivation curves to higher temperatures. However, at 25°C the kcat of P. lapideum AspAT was nearly equal to the values of AspATs from mesophilic organisms. The enzyme used L-aspartate and L-cysteine sulfinate as amino donors and 2-oxoglutarate as an amino acceptor. The K m values were 5.0 mM for L-aspartate, 5.7 mM for L-glutamate, 0.2 mM for 2-oxoglutarate, and 0.032 mM for oxaloacetate. © 2003 by Japan Society for Bioscience, Biotechnology, and Agrochemistry.