PURIFICATION AND PARTIAL CHARACTERIZATION OF BARLEY GLUTAMYL-TRNA(GLU) REDUCTASE, THE ENZYME THAT DIRECTS GLUTAMATE TO CHLOROPHYLL BIOSYNTHESIS

5-Aminolevulinic acid for chlorophyll synthesis in greening barley is formed from glutamate. One of the steps involved in the conversion of glutamate to 5-aminolevulinic acid involves a reduction of glutamyl-tR NA(Glu) to glutamate 1-semialdehyde and tRNA(Glu). An enzyme catalysin g this reduction was purified from the stroma of greening barley chlor oplasts. An approximately 270-kDa protein composed of 54-kDa identical subunits was identified as the barley glutamyl tRNA(Glu) reductase af ter purification by Sephacryl S-300, Cibacron Blue-Sepharose, 2'-5'-AD P-Sepharose, Mono S, Mini Q and Superose 12 chromatography. The sequen ce of 18 amino acids from the N-terminus of the reductase is 50% ident ical to a cDNA-deduced domain of the Arabidopsis thaliana hemA protein and encoded in a barley hemA cDNA sequence. This is an unequivocal de monstration that the glutamyl-tRNA(Glu) reductase subunit of higher pl ants is encoded in a hemA gene of the nuclear genome. Heme at 4 mu M c oncentration or glutamate 1-semialdehyde at 200 mu M caused a 50% inhi bition of the reductase activity. Micromolar concentrations of Zn2+, C u2+ and Cd2+ also inhibited barley glutamyl-tRNA(Glu) reductase.