Abstract
We have recently found that the glutathione-S-transferase π-isozyme (GST-π), a cellular detoxification enzyme, potently and selectively inhibits activation of jun protein by its upstream kinase, jun kinase (JNK). This newly identified regulatory activity of GST-π is strongly inhibited by a group of agents that inhibit its enzymatic activity. Since loss of enzymatic activity in general does not correlate with loss of regulatory activity, it is likely that inhibitor binding induces changes in the structure of one or more domains of GST that block its interaction with JNK. To identify regions of GST that change conformation on the binding of inhibitors, we have performed molecular dynamics calculations on GST-π to compute its average structure in the presence and absence of the inhibitor, glutathione sulfonate. Superposition of the two average structures reveals that several regions change local structure depending upon whether the inhibitor is bound or not bound. Two of these regions, residues 36–50 and 194–201, are highly exposed. We have synthesized peptides corresponding to these two segments and find that the 194–201 sequence strongly inhibits the ability of GST-π to block the in vitro phosphorylation of jun by JNK. These results suggest that this region of GST-π is critical to its functioning as a newly discovered regulator of signal transduction.
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REFERENCES
Adler, V., Pincus, M. R., Brandt-Rauf, P. W., and Ronai, Z. (1995). Complexes of ras-p21 with jun-N-Kinase and c-jun Proteins, Proc. Natl. Acad. Sci. USA 92, 10585–10589.
Adler, V., Pincus, M. R., Polatskaya, A., Montano, X., Friedman, F. K., and Ronai, Z. (1996). Activation of c-jun NH2 kinase by UV irradiation is dependent on p21ras. J. Biol. Chem. 271, 23304–23309.
Adler, V., Yin, Z., Fuchs, S., Benezra, M., Rosario, L., Tew, K., Pincus, M. R., Sardana, M., Henderson, C., Wolf, C. R., Davis, R., and Ronai, Z. (1999). GSTπ—A regulator of JNK signaling, EMBO J. 18, 1321–1334.
Amar, S., Golozman, A., Chung, D. L., Alder, V., Ronai, Z., Friedman, F. K., Robinson, R., Brandt-Rauf, P. W., Yamaizumi, Z., and Pincus, M. R. (1997). Selective inhibition of oncogenic ras-p21 in vivo by agents that block its interaction with jun-N-kinase (JNK) and jun proteins. Implications for the design of selective chemotherapeutic agents. Cancer Chemother. Pharmacol. 41, 79–85.
Chen, J. M., Manolatos, S., Brandt-Rauf, P. W., Monaco, R., and Pincus, M. R. (1996). Computed three-dimensional structures for the ras-binding domain of the raf-p74 protein complexed with ras-p21 and with its suppressor protein, rap-1A. J. Protein Chem. 15, 511–518.
Dauber-Osguthorpe, P., Roberts, V. A., Osguthorpe, D. J., Wolff, J., Genest, M., and Hagler, A. T. (1988). Structure and energetics of ligand binding to proteins: E. coli dihydrofolate reductase-trimethroprim, a drug-receptor system. Proteins Struct. Funct. Genet. 4, 31–47.
Deshpande, A. K., and Kung, H.-F. (1987). Insulin induction of Xenopus laevis oocyte maturation is inhibited by monoclonal antibody against p21 ras proteins, Mol. Cell. Biol. 7, 1285–1288.
Flatgaard, J. E., Bauer, K. E., and Kauvar, K. M. (1993) Isozyme specificity of novel glutathione-S-Transferase inhibitors. Cancer Pharmacol. Chemother. 33, 63–70.
Garcia-Saez, I., Parraga, A., Phillips, M. F., Mantle, T. J., and Coll, M. (1994), Molecular structure at 1.8 Å of mouse liver class pi glutathione-S-transferase complexed with S-(p-nitrobenzyl)glutathione and other inhibitors, J. Mol. Biol. 237, 298–314.
Kantor, R., Eschenfelder, P., and Pincus, M. R. (1991a). The piisozyme of glutathione S-transferase may be a serum marker for solid tissue (epithelial cell) tumors. In American Association for Clinical Chemistry, 43rd National Meeting, Washington, D.C., p. 680.
Kantor, R. R. S., Giardina, S. L., Bartolazzi, A., Townsend, A. J., Myers, C. E., Cowan, K. H., Longo, D. L., and Natali, P. G. (1991b). Monoclonal antibodies to glutathione-S-transferase π-immunochemical analysis of human tissues and cancers. Int. J. Cancer 47, 193–201.
Liwo, A., Gibson, K. D., Scheraga, H. A., Brandt-Rauf, P. W., Monaco, R., and Pincus, M. R. (1994). Comparison of the low energy conformations of an oncogenic and a non-oncogenic p21 protein, neither of which binds GDP or GTP, J. Protein Chem. 13, 237–251.
Mannervik, B., and Danielson, H. U. (1988). Glutathione transferases–Structure and catalytic activity, CRC. Crit. Rev. Biochem. 23, 283–337.
Meyer, D. J., Coles, B., Pemble, S. E., Glimore, K. S., Fraser, G. M., and Ketterer, B. (1991). Theta, a new class of glutathione transferases purified from rat and man, Biochem. J. 274, 409–414.
Monaco, R., Chen, J. M., Chung, D. L., Brandt-Rauf, P. W., and Pincus, M. R. (1995a). Comparison of the computed three-dimensional structures of oncogenic forms of the ras-gene-encoded p21 protein with the structure of the normal (non-transforming) wild-type protein, J. Protein Chem. 14, 457–466.
Monaco, R., Chen, J. M., Friedman, F. K., Brandt-Rauf, P. W., and Pincus, M. R. (1995b). Structural effects of the binding of GTP to the wild-type and oncogenic forms of the ras-gene-encoded p21 proteins, J. Protein Chem. 14, 721–730.
Nemethy, G., Pottle, M. S., and Scheraga, H. A. (1983). Energy parameters in polypeptides. 9. Updating of geometrical parameters, nonbonded interactions and hydrogen bond interactions for the naturally occurring amino acids, J. Phys. Chem. 87, 1883–1887.
Pincus, M. R., Brandt-Rauf, P. W., Michl, J., and Friedman, F. K. (2000). ras-p21-induced cell transformation: Unique signal transduction pathways and implications for the design of new chemotherapeutic agents, Cancer Invest., 18, 39–50.
Post, C. B., Brooks, B. R., Karplus, M., Dobson, C. M., Artymiuk, P. J., Cheetham, J. C., and Phillips, D. C. (1986). Molecular dynamics simulations of native and substrate-bound lysozyme. A study of the average structures and atomic fluctuations, J. Mol. Biol. 190, 455–479.
Villafania, A., Anwar, K., Amar, S., Chung, D., Adler, V., Ronai, Z., Brandt-Rauf, P. W., Yamaizumi, Z., Kung, H.-F., and Pincus, M. R. (2000). Glutathione-S-transferase as a selective inhibitor of oncogenic ras-p21-induced mitogenic signaling through blockade of activation of jun by jun-N-terminal kinase, Ann. Clin. Lab. Sci., 30, 57–64.
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Monaco, R., Friedman, F.K., Hyde, M.J. et al. Identification of a Glutathione-S-Transferase Effector Domain for Inhibition of jun Kinase, by Molecular Dynamics. J Protein Chem 18, 859–866 (1999). https://doi.org/10.1023/A:1020679229110
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DOI: https://doi.org/10.1023/A:1020679229110