Subunit interaction in unadenylylated glutamine synthetase from Escherichia coli. Evidence from methionine sulfoximine inhibition studies.
Although glutamine synthetase from Escherichia coli is composed of 12 identical subunits, there is no evidence that homologous subunit interactions occur in fully unadenylylated or fully adenylylated enzyme. Meister and co-workers (Manning, J. M., Moore, S., Rowe, W. B., and Meister, A. (1969) Biochemistry 8, 2681-2685) have shown that L-methionine-S-sulfoximine, one of the four diastereomers of methionine sulfoximine, preferentially inhibits glutamine synthetase irreversibly in the presence of ATP, due to the formation of tightly bound products, ADP, and methionine sulfoximine phosphate. Using highly purified unadenylylated glutamine synthetase and the two resolved diastereomers of L-methionine-S,R-sulfoximine, we have studied both the kinetics of glutamine synthetase inactivation in the presence of excess methionine sulfoximine and ATP, and the binding of methionine sulfoximine to the enzyme. The results reveal that (a) the apparent first order rate constant of irreversible inactivation by the S isomer decreases progressively from the expected first order rate, indicating that an inactivated subunit retards the reactivity of its neighboring subunits toward methionine sulfoximine and ATP; (b) the R isomer does not inactivate glutamine synthetase irreversibly in the presence of ATP; however, the R isomer is capable of protecting the enzyme temporarily from the irreversible inhibition by the S isomer; and (c) the binding of the S isomer monitored by changes in protein fluorescence exhibits an apparent negative cooperative binding isotherm, whereas the R isomer yields an apparent positive cooperative pattern.
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D005974:Glutamate-Ammonia Ligase / Q000378:metabolism*; D007536:Isomerism; D007700:Kinetics; D046911:Macromolecular Substances; D008717:Methionine Sulfoximine / Q000494:pharmacology*; D011485:Protein Binding; D013050:Spectrometry, Fluorescence
D046911:Macromolecular Substances; D008717:Methionine Sulfoximine; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D005974:Glutamate-Ammonia Ligase