[Presence of SH-groups and histidine in the active site of Chlorella glutamine synthetase].
The presence of two cysteine residues per each six monomers comprising the oligomer of Chlorella glutamine synthetase (E.C.6.3.1.2) is demonstrated using homogenous enzyme preparation. p-Chloromercuribenzoate (p-CMB) is found to inhibit glutamine synthetase activity, the degree of inhibition depending on the inhibitor concentration. The following enzyme reactivation by dithiotreitol (10(-2) M) was observed only when the enzyme was inactivated with 10(-5) M p-CMB under 15 min. preincubation. Preincubation of the enzyme with 10(-4) M p-CMB for 45 min. did not result in its reactivation. Gel filtration of glutamine synthetase treated with 10(-4) M p-CMB has revealed the dissociation of the enzyme into inactive monomers. Incubation of glutamine synthetase with p-CMB at various pH values, incubation after pre-treatment with urea and experiments with HgCl2 indicate the presence of free and masked inside the globula SH-groups in the enzyme molecule. Competitive character of the enzyme inhibition with p-CMB with respect to ATP indicates that SH-groups of the active site participate in the ATP binding, probably, as Mg-ATP or Mn-ATP complexes. Data on the estimation of ionization constant of glutamate-binding group and experiments on the effect of histidine photooxidation on the enzyme activity indicate the presence of histidine residue in the enzyme active site, which participates in glutamate binding.
D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites; D002708:Chlorella / Q000201:enzymology*; D002729:Chloromercuribenzoates / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D004793:Enzyme Reactivators; D005974:Glutamate-Ammonia Ligase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006639:Histidine / Q000378:metabolism*; D046911:Macromolecular Substances; D011485:Protein Binding; D013438:Sulfhydryl Compounds / Q000378:metabolism*
D002729:Chloromercuribenzoates; D004793:Enzyme Reactivators; D046911:Macromolecular Substances; D013438:Sulfhydryl Compounds; D006639:Histidine; D000255:Adenosine Triphosphate; D005974:Glutamate-Ammonia Ligase; D004229:Dithiothreitol
O nalichii SH-Grupp i gistidina v aktivnom tsentre glutaminsintetazy kholerally.