Purification and partial characterization of glutamine synthetase isoforms from Triticale seedlings.

Abstract

Two isoforms of glutamine synthetase (EC 6.3.1.2), cytoplasmic (GS1) and chloroplastic (GS2) were isolated from shoots of 14-day-old Triticale seedlings, and purified 260-fold and 248-fold, respectively. Specific activities of the two preparations were 35.1 and 33.5 mumol x min-1 per mg of protein, respectively. Both crude extracts and homogeneous GS1 and GS2 preparations required divalent metal ions (Mg2+, Mn2+, Co2+) for their activities. Mg2+ was the most effective activator, the highest activity of GS1 being reached at 5 mM, and that of GS2 at 20 mM MgCl2. The optimum pH for the two isoforms showed large differences, dependent also on the kind of divalent ion. Molecular masses of GS1 and GS2 were 305000 Da and 385200 Da, respectively. It seems that native protein of GS1 is built from eight identical subunits of Mm 38000 Da and that of GS2 of the same number of subunits but of Mm of about 48000 Da. Proteins of GS isoforms differed significantly in their amino-acid composition.