Characterization of aspartate aminotransferase from the cyanobacterium Phormidium lapideum.
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 Ialpha of AspATs from eukaryotes and Escherichia coli, but it was similar to sequences of the subfamily Igamma of AspATs from archaebacteria and eubacteria. The enzyme was most active at 80 degrees C and was stable at up to 75 degrees C. Thermal inactivation (60-85 degrees 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 degrees 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 Km 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.
Kim
Hyeung
H
Department of Life Science and Biotechnology, Faculty of Life and Environmental Science, Shimane University, 1060 Nishikawatsu, Matsue, Shimane 690-8504, Japan.
Ikegami
Koji
K
Nakaoka
Masaki
M
Yagi
Mayumi
M
Shibata
Hitoshi
H
Sawa
Yoshihiro
Y
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001219:Aspartate Aminotransferases / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D001224:Aspartic Acid / Q000378:metabolism; D002384:Catalysis; D000458:Cyanobacteria / Q000201:enzymology*; D003545:Cysteine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007656:Ketoglutaric Acids / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D018377:Neurotransmitter Agents; D017386:Sequence Homology, Amino Acid
D000596:Amino Acids; D007656:Ketoglutaric Acids; D018377:Neurotransmitter Agents; D001224:Aspartic Acid; D001219:Aspartate Aminotransferases; D003545:Cysteine; C013461:cysteine sulfinic acid