Aspartate aminotransferase isozymes in Panicum miliaceum L., an NAD-malic enzyme-type C4 plant: comparison of enzymatic properties primary structures, and expression patterns.
Panicum miliaceum has at least three isozymes of aspartate aminotransferase (AspAT); the cytosolic and mitochondrial isozymes (cAspAT and mAspAT) are major components and the third is a minor isozyme. Fractionation of leaf subcellular components showed that the minor isozyme was localized in plastids (pAspAT). We purified the three isozymes from green leaves of P. miliaceum. Both cAspAT and pAspAT consisted of triple subforms having the same molecular size but different isoelectric points. No substantial difference in enzymatic properties was observed among these isozymes besides the pH profiles. We isolated a full-length cDNA clone for pAspAT. This clone contains an open reading frame that encodes 457 amino acids. The amino-terminal region of the pAspAT precursor shares common features of plastid transit peptides. The amino acid sequence of P. miliaceum pAspAT shows higher similarity with other plant pAspATs than P. miliaceum cAspAT and mAspAT. The mRNA levels of the three isozymes were high in leaves compared with roots and mesocotyls. The three isozymes showed different expression patterns against environmental stimuli such as light and nitrate. The activities and protein levels of cAspAT and mAspAT increased during greening in accordance with those of phosphoenolpyruvate carboxylase and NAD-malic enzyme involved in the C4 pathway, primarily as a consequence of the increase in the levels of their mRNAs. By contrast, pAspAT was constitutively expressed during greening. The activity and protein levels of cAspAT and mAspAT selectively increased during recovery from an nitrogen deficit, primarily as a consequence of increase in the levels of their mRNAs while those of pAspAT remained unchanged.
Taniguchi
M
M
Department of Applied Biological Sciences, School of Agricultural Sciences, Nagoya University, Japan.
Kobe
A
A
Kato
M
M
Sugiyama
T
T
D000595:Amino Acid Sequence; D001219:Aspartate Aminotransferases / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D018076:DNA, Complementary; D017886:Durapatite; D015870:Gene Expression; D007527:Isoenzymes / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018515:Plant Leaves; D010944:Plants / Q000201:enzymology*; D016133:Polymerase Chain Reaction
D018076:DNA, Complementary; D007527:Isoenzymes; D017886:Durapatite; D001219:Aspartate Aminotransferases