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Mg2+ is bound to glutamine synthetase extracted from bovine or ovine brain in the presence of l-Methionine-S-sulfoximine phosphate

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Abstract

Purified glutamine synthetase from bovine or ovine brain had no tightly bound Mn2+. By extraction of bovine or ovine brain glutamine synthetase in the presence of l-Met-S-sulfoximine phosphate and ADP in metal ion-free water and 0.1 m KCl, only endogenously bound divalent cations were trapped on the enzyme. Enzyme complexes isolated by immunoprecipitation contained <0.05 Mn2+ and 1.5 ± 0.2 Mg2+ per subunit. Without inactive complex formation, the enzyme immunoprecipitated from extracts contained undetectable Mn2+ (<0.01 eq per subunit) and 0.1–2.0 eq of Mg2+ per subunit. Direct binding measurements showed that the purified bovine brain enzyme contained two divalent cations bound at the active site of each subunit. Thus, although either Mg2+ or Mn2+ supports enzyme activity in vitro, Mg2+ rather than Mn2+ appears to be bound to brain glutamine synthetase in vivo.

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      2000, Biochimica et Biophysica Acta - Protein Structure and Molecular Enzymology
      Citation Excerpt :

      Most of the biochemical properties have been established using enzyme derived from sheep brain [67]. Forty years of studies of the enzymology of eukaryotic GS have included work on the following tissues: rat liver [68–70], rabbit and rat skeletal muscle [71], pig brain [72,73], bovine brain [74–76] and human brain [77–79], as well as many studies on sheep brain [61,67,75,80–87]. For reviews on eukaryotic GS see Meister [71], Wedler and Denman [88], Wedler and Toms [4] and Purich [5].

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    Present address: Laboratory of Molecular Biology, National Cancer Institute, Building 37, Room 2E20, Bethesda, Md. 20892.

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