Sa. Mccallum et al., Solution structure of the carboxyl terminus of a human class Mu glutathione S-transferase: NMR assignment strategies in large proteins, J MOL BIOL, 285(5), 1999, pp. 2119-2132
Strategies to obtain the NMR assignments for the H-N, N, CO, C-alpha and C-
beta resonance frequencies for the human class mu glutathione-S-transferase
GSTM2-2 are reported. These assignments were obtained with deuterated prot
ein using a combination of scalar and dipolar connectivities and various sp
ecific labeling schemes. The large size of this protein (55 kDa, homodimer)
necessitated the development of a novel pulse sequence and specific labeli
ng strategies. These aided in the identification of residue type and were e
ssential components in determining sequence specific assignments. These ass
ignments were utilized in this study to characterize the structure and dyna
mics of the carboxy-terminal residues in the unliganded protein. Previous c
rystallographic studies of this enzyme in complex with glutathione suggeste
d that this region may be disordered, and that this disorder may be essenti
al for catalysis. Furthermore, in the related class alpha protein extensive
changes in conformation of the C terminus are observed upon ligand binding
. On the basis of the results presented here, the time-averaged conformatio
n of the carboxyl terminus of unliganded GSTM2-2 is similar to that seen in
the crystal structure. NOE patterns and H-1-N-15 heteronuclear nuclear Ove
rhauser enhancements suggest that this region of the enzyme does not underg
o motion on a rapid time scale. (C) 1999 Academic Press.