Conformational differences between unadenylylated and adenylylated glutamine synthetase from Escherichia coli on binding L-methionine sulfoximine.

Abstract

L-Methionine-SR-sulfoximine (a proposed transition state analog for glutamine synthetase) produces tyrosyl residue perturbations on binding to unadenylylated but not to adenylylated glutamine synthetase from Escherichia coli, as evidenced by UV difference spectra.Binding L-methionine-SR-sulfoximine to complexes of unadenylylated glutamine synthetase (manganese enzyme, magnesium * enzyme, and ADP.manganese-enzyme) produces red-shifted, UV difference spectra consistent with the burial of approximately 1 tyrosyl residue per unadenylylated subunit.This may involve primarily the tyrosyl residue at the subunit site of adenylylation because the binding of L-methionine-SR-sulfoximine to the fully adenylylated manganese enzyme produces a red-shifted spectral perturbation of covalently bound 5"adenylate groups without a concomitant tyrosyl perturbation.Spectrophotometric results indicate also that both L-methionine-S-sulfoximine and L-methionine-R-sulfoximine in a mixture of the two isomers bind reversibly to glutamine synthetase and compete for a single subunit site.Hill plots of spectrophotometric titrations with L-methionine-SR-sulfoximine (treating the diastereoisomeric mixture as a single ligand) give for the unadenylylated enzyme: [SI., = 46 PM and nH = 0.9 for manganese=enzyme; [S]O.S = 9 PM and RH = 1.0 for ADP manganese enzyme complex; and [SIo., = 1.9 m M and nH = 0.5 for magnesium enzyme.For binding L-methionine-SR-sulfoximine to the fully adenylylated manganese enzyme, [ao.5 = 0.19 m~ and