Noncovalent associations of glutathione S-transferase and ligands: A study using electrospray quadrupole/time-of-flight mass spectrometry
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Human glutathione S-transferase A1-1 was observed predominantly as dimeric ions (51 kDa) during electrospray mass spectrometric analysis from aqueous solution at pH 7.4, in keeping with the known dimeric structure in solution. When analyses were performed on solutions of the enzyme containing glutathione (GSH), noncovalent adducts of protein dimer and one or two ligand molecules were observed; each mass increment, which exceeded the mass of GSH alone, was provisionally interpreted to indicate concomitant association of two water molecules per bound GSH. Noncovalent adducts of ligand and protein dimer were similarly observed for oxidized glutathione and for two glutathione inhibitors, both incorporating substituted thiol structures. In these instances, the mass increments exactly matched the ligand masses, suggesting that the apparent concomitant binding of water was associated with the presence in the ligand of a free thiol group. Collisionally activated decomposition during tandem mass spectrometry analyses of noncovalent adducts incorporating protein dimer and ligands yielded initially the denuded dimer; at higher collision energies the monomer and a protein fragment were formed.
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- 11.Murayama, K.; Taka, H.; Kaga, N.; Fujimura, T.; Mineki, R.; Shindo, N.; Morita, M.; Hosono, M.; Nitta, K. The structure of Silurus asotus (catfish) roe lectin (SAL): Identification of a noncovalent trimer by mass spectrometry and analytical ultracentrifugation. Anal. Biochem. 1997, 247, 319–326.CrossRefGoogle Scholar
- 19.Armstrong, R. N. Glutathione S-transferase—structure and mechanism of an archetypical detoxication enzyme. Adv. Enzymol. Relat. Areas Mol. Biol. 1994, 69, 1–44.Google Scholar
- 21.Mannervik, B. The isozymes of glutathione transferase. Adv. Enzymol. Relat. Areas Mol. Biol. 1985, 57, 357–417.Google Scholar
- 22.van Ommen, B.; Bogaards, J. J.; Peters, W. H.; Blaauboer, B.; Van Bladeren, P. J. Quantification of human hepatic glutathione S-transferases. Biochem. J. 1990, 269, 609–613.Google Scholar
- 23.Sinning, I.; Kleywegt, G. J.; Cowan, S. W.; Reinemer, P.; Dirr, H. W.; Huber, R.; Gilliand, G. L.; Armstrong, R. N.; Ji, X.; Board, P. G.; Olin, B.; Mannervik, B.; Jones, T. A. Structure determination and refinement of human alpha class glutathione transferase A1-1, and a comparison with the mu and pi class enzymes. J. Mol. Biol. 1993, 232, 192–212.CrossRefGoogle Scholar
- 24.Camereon, A. D.; Sinning, I.; L’Hermite, G.; Olin, B.; Board, P. G.; Mannervik, B.; Jones, T. A. Structural analysis of human alpha-class glutathione transferase A1-1 ion the apo-form and in complexes with ethacrynic acid and its glutathione conjugate. Structure 1995, 3, 717–727.CrossRefGoogle Scholar
- 28.Cooke, R. J.; Bjornestedt, R.; Douglas, K. T.; McKie, J. H.; King, M. D.; Coles, B.; Ketterer, B.; Mannervik, B. Photoaffinity labelling of the active site of the rat glutathione transferases 3-3 and 1-1 and human glutathione transferase A1-1. Biochem. J. 1994, 302, 383–390.Google Scholar
- 36.Chung, E.; Henriques, D.; Renzoni, D.; Zvelebil, M.; Bradshaw, J. M.; Waksman, G.; Robinson, C. V.; Ladbury, J. E. Mass spectrometric and thermodynamic studies reveal the role of water molecules in complexes formed between SH2 domains and tyrosyl phosphopeptides. Structure 1998, 6, 1141–1151.CrossRefGoogle Scholar