JBPC  Vol.3 No.3 , August 2012
Homology modeling and structural analysis of human γ-glutamylcysteine ligase catalytic subunit for antitumor drug development
ABSTRACT
Homology modeling and structural analysis of human glutamate cysteine ligase catalytic subunit (hGCLC) were performed with a software package the Molecular Operating Environment. A yeast GCLC (yGCLC; PDB code: 3LVV) was selected as a template for the 3D structure modeling of hGCLC. The modeled hGCLC showed significant 3D similarities at the ligand biding site (LBS) to the yGCLC structure. The contact energy profiles of the hGCLC model were in good agreement with those of the yGCLC structure. Ramachandran plots revealed that only 1.4% of the amino acid residues were in the disfavored region for hGCLC. The molecular electrostatic potential (MEP) map of the hGCLC model exhibited that the model was slightly different from the yGCLC model electrostatically at the LBS. Further, docking simulations revealed the similarity of the ligand-receptor bound location between the hGCLC and yGCLC models. The different binding orientations between the glutathione (GSH)-hGCLC and GSH-yGCLC complexes reflected the different MEP maps at the LBSs between the hGCLC and yGCLC models. These results indicate that the hGCLC model was successfully modeled and analyzed. To the best of our knowledge, this is the first report of a hGCLC model with detailed analyses, and our data verify that the model can be utilized for application to target hGCLC for the development of anticancer drugs.

Cite this paper
Yamaguchi, H. , Akitaya, T. , Kidachi, Y. , Kamiie, K. and Umetsu, H. (2012) Homology modeling and structural analysis of human γ-glutamylcysteine ligase catalytic subunit for antitumor drug development. Journal of Biophysical Chemistry, 3, 238-248. doi: 10.4236/jbpc.2012.33028.
References
[1]   Griffith, O.W. and Mulcahy, R.T. (1999) The enzymes of glutathione synthesis: gamma-glutamylcysteine synthetase. Advances in Enzymology and Related Areas of Molecular Biology, 73, 209-267. doi:10.1002/9780470123195.ch7

[2]   Orlowski, M. and Meister, A. (1971) Partial reactions catalyzed by γ-glutamylcysteine synthetase and evidence for an activated glutamate intermediate. The Journal of Biological Chemistry, 246, 7095-7105.

[3]   Strumeyer, D.H. and Bloch, K. (1960) Some properties of gamma-glutamylcysteine synthetase. The Journal of Biological Chemistry, 235, PC27.

[4]   Yip, B. and Rudolph, F.B. (1976) The kinetic mechanism of rat kidney gamma-glutamylcysteine synthetase. The Journal of Biological Chemistry, 251, 3563-3568.

[5]   Meister, A. and Anderson, M.E. (1983) Glutathione. Annual Review of Biochemistry, 52, 711-760. doi:10.1146/annurev.bi.52.070183.003431

[6]   Forman, H.J., Liu, R. and Shi, M.M. (1995) Glutathione synthesis in oxidative stress. In: Packer, L. and Cadenas, E., Eds., Biothiols in Health and Disease, Marcel Dekker, New York, 189-212.

[7]   Hall, A.G. (1999) The role of glutathione in the regulation of apoptosis. European Journal of Clinical Investigation, 29, 238-245. doi:10.1046/j.1365-2362.1999.00447.x

[8]   Meister, A. (1991) Glutathione deficiency produced by inhibition of its synthesis, and its reversal; applications in research and therapy. Pharmacology & Therapeutics, 51, 155-194. doi:10.1016/0163-7258(91)90076-X

[9]   Manna, S.K., Kuo, M.T. and Aggarwal, B.B. (1999) Over- expression of gamma-glutamylcysteine synthetase suppresses tumor necrosis factor-induced apoptosis and activation of nuclear transcription factor-kappa B and activator protein-1. Oncogene, 18, 4371-4382. doi:10.1038/sj.onc.1202811

[10]   Botta, D., Franklin, C.C., White, C.C., Krejsa, C.M., Dabrowski, M.J., Pierce, R.H., Fausto, N. and Kavanagh, T.J. (2004) Glutamate-cysteine ligase attenuates TNF-induced mitochondrial injury and apoptosis. Free Radical Biology & Medicine, 37, 632-642. doi:10.1016/j.freeradbiomed.2004.05.027

[11]   Tateishi, N., Higashi, T., Shinya, S., Naruse, A. and Sakamoto, Y. (1974) Studies on the regulation of glutathione level in rat liver. The Journal of Biological Chemistry, 75, 93-103.

[12]   Wild, A.C. and Mulcahy, R.T. (1999) Pyrrolidine dithiocarbamate up-regulates the expression of the genes encoding the catalytic and regulatory subunits of gammaglutamylcysteine synthetase and increases intracellular glutathione levels. Biochemical Journal, 338, 659-665. doi:10.1042/0264-6021:3380659

[13]   Lu, S.C. (2009) Regulation of glutathione synthesis. Molecular Aspects of Medicine, 30, 42-59. doi:10.1016/j.mam.2008.05.005

[14]   Godwin, A.K., Meister, A., O’Dwyer, P.J., Huang, C.S., Hamilton, T.C. and Anderson, M.E. (1992) High resistance to cisplatin in human ovarian cancer cell lines is associated with marked increase of glutathione synthesis. Proceedings of the National Academy of Sciences of the United States of America, 89, 3070-3074. doi:10.1073/pnas.89.7.3070

[15]   Mulcahy, R.T., Bailey, H.H. and Gipp, J.J. (1994) Up-regulation of gamma-glutamylcysteine synthetase activity in melphalan-resistant human multiple myeloma cells expressing increased glutathione levels. Cancer Chemotherapy and Pharmacology, 34, 67-71. doi:10.1007/BF00686114

[16]   Mulcahy, R.T., Bailey, H.H. and Gipp, J.J. (1995) Transfection of complementary DNAs for the heavy and light subunits of human gamma-glutamylcysteine synthetase results in an elevation of intracellular glutathione and resistance to melphalan. Cancer Research, 55, 4771-4775.

[17]   Anderson, M.E. (1998) Glutathione: An overview of biosynthesis and modulation. Chemico-Biological Interactions, 111-112, 1-14. doi:10.1016/S0009-2797(97)00146-4

[18]   Townsend, D.M. and Tew, K.D. (2003) The role of glutathione-S-transferase in anti-cancer drug resistance. Oncogene, 22, 7369-7375. doi:10.1038/sj.onc.1206940

[19]   Griffith, O.W. (1982) The Journal of Biological Chemistry, 257, 13704-13712.

[20]   Yamaguchi, H., Yu, T., Noshita, T., Kidachi, Y., Kamiie, K., Yoshida, K., Akitaya, T., Umetsu, H. and Ryoyama, K. (2011) Ligand-receptor interaction between triterpenoids and the 11beta-hydroxysteroid dehydrogenase type 2 (11-betaHSD2) enzyme predicts their toxic effects against tumorigenic r/m HM-SFME-1 cells. The Journal of Biological Chemistry, 286, 36888-36897. doi:10.1074/jbc.M111.265900

[21]   Seelig, G.F. and Meister, A. (1984) Gamma-glutamylcys-teine synthetase. Interactions of an essential sulfhydryl group. The Journal of Biological Chemistry, 259, 3534-3538.

[22]   Biterova, E.I. and Barycki, J.J. (2009) Mechanistic details of glutathione biosynthesis revealed by crystal structures of Saccharomyces cerevisiae glutamate cysteine ligase. The Journal of Biological Chemistry, 284, 32700-32708. doi:10.1074/jbc.M109.025114

[23]   Biterova, E.I. and Barycki, J.J. (2010) Structural basis for feedback and pharmacological inhibition of Saccharomyces cerevisiae glutamate cysteine ligase. The Journal of Biological Chemistry, 285, 14459-14466. doi:10.1074/jbc.M110.104802

[24]   Kurogi, Y. and Guner, O.F. (2001) Pharmacophore modeling and three-dimensional database searching for drug design using catalyst. Current Medicinal Chemistry, 8, 1035-1055.

[25]   Ekins, S. (2004) Predicting undesirable drug interactions with promiscuous proteins in silico. Drug Discovery Today, 9, 276-285. doi:10.1016/S1359-6446(03)03008-3

[26]   Jorgensen, W.L. (2004) The many roles of computation in drug discovery. Science, 303, 1813-1818. doi:10.1126/science.1096361

[27]   Yamaguchi, H., Akitaya, T., Yu, T., Kidachi, Y., Kamiie, K., Noshita, T., Umetsu, H. and Ryoyama, K. (2011) Homology modeling and structural analysis of 11β-hydroxysteroid dehydrogenase type 2. European Journal of Medicinal Chemistry, 46, 1325-1330. doi:10.1016/j.ejmech.2011.01.054

[28]   Le, T.M., Willis, A.S., Barr, F.E., Cunningham, G.R., Canter, J.A., Owens, S.E., Apple, R.K., Ayodo, G., Reich, D. and Summar, M.L. (2010) An ethnic-specific polymorphism in the catalytic subunit of glutamate-cysteine ligase impairs the production of glutathione intermediates in vitro. Molecular Genetics and Metabolism, 101, 55-61. doi:10.1016/j.ymgme.2010.05.013

[29]   Levitt, M. (1992) Accurate modeling of protein conformation by automatic segment matching. Journal of Molecular Biology, 226, 507-533. doi:10.1016/0022-2836(92)90964-L

[30]   Fechteler, T., Dengler, U. and Schomberg, D. (1995) Prediction of protein three-dimensional structures in insertion and deletion regions: a procedure for searching data bases of representative protein fragments using geometric scoring criteria. Journal of Molecular Biology, 253, 114-131. doi:10.1006/jmbi.1995.0540

[31]   Zhang, C., Vasmatizis, G., Cornette, J.L. and DeLisi, C. (1997) Determination of atomic desolvation energies from the structures of crystallized proteins. Journal of Molecular Biology, 267, 707-726. doi:10.1006/jmbi.1996.0859

[32]   Bowie, J.U., Lüthy, R. and Eisenberg, D. (1991) A method to identify protein sequences that fold into a known three-dimensional structure. Science, 253, 164-170. doi:10.1126/science.1853201

[33]   Lüthy, R., Bowie, J.U. and Eisenberg, D. (1992) Assessment of protein models with three dimensional profiles. Nature, 356, 83-85. doi:10.1038/356083a0

[34]   Carvajal, C.A., Gonzalez, A.A., Romero, D.G., González, A., Mosso, L.M., Lagos, E.T., Hevia, Mdel.P., Rosati, M.P., Perez-Acle, T.O., Gomez-Sanchez, C.E., Montero, J.A. and Fardella, C.E. (2003) Two homozygous mutations in the 11 beta-hydroxysteroid dehydrogenase type 2 gene in a case of apparent mineralocorticoid excess. The Journal of Clinical Endocrinology and Metabolism, 88, 2501-2507. doi:10.1210/jc.2002-021909

[35]   Liang, J., Edelsbrunner, H., Fu, P., Sudhakar, P.V. and Subramaniam, S. (1998) Analytical shape computation of macromolecules: I. Molecular area and volume through alpha shape. Proteins, 33, 1-17.

[36]   Liang, J., Edelsbrunner, H., Fu, P., Sudhakar, P.V. and Subramaniam, S. (1998) Analytical shape computation of macromolecules: II. Inaccessible cavities in proteins. Proteins, 33, 18-29.

[37]   Goto, J., Kataoka, R. and Hirayama, N. (2004) Ph4Dock: pharmacophore-based protein-ligand docking. Journal of Medicinal Chemistry, 47, 6804-6811. doi:10.1021/jm0493818

[38]   Goto, J., Kataoka, R., Muta, H. and Hirayama, N. (2008) ASEDock-docking based on alpha spheres and excluded volumes. Journal of Chemical Information and Modeling, 48, 583-590. doi:10.1021/ci700352q

[39]   Halgren, T.A. (1996) Merck molecular force field. I. basis, form, scope, parameterization and performance of MM-FF94. Journal of Computational Chemistry, 17, 490-519.

[40]   Abbott, J.J., Pei, J., Ford, J.L., Qi, Y., Grishin, V.N., Pitcher, L.A., Phillips, M.A. and Grishin, N.V. (2001) Structure prediction and active site analysis of the metal binding determinants in gamma-glutamylcysteine synthetase. The Journal of Biological Chemistry, 276, 42099-42107. doi:10.1074/jbc.M104672200

[41]   Lovell, S.C., Davis, I.W., Arendall III, W.B., de Bakker, P.I., Word, J.M. and Prisant, M.G., Richardson, J.S. and Richardson, D.C. (2003) Structure validation by Cα geometry: φ, ψ and Cβ deviation, Proteins, 50, 437-450. doi:10.1002/prot.10286

[42]   Hibi, T., Nii, H., Nakatsu, T., Kimura, A., Kato, H., Hiratake, J. and Oda, J. (2004) Crystal structure of γ-glutamylcysteine synthetase: Insights into the mechanism of catalysis by a key enzyme for glutathione homeostasis. Proceedings of the National Academy of Sciences of the United States of America, 101, 15052-15057. doi:10.1073/pnas.0403277101

[43]   Metropolis, N. and Ulam, S. (1949) The Monte Carlo method. Journal of the American Statistical Association, 44, 335-341. doi:10.1080/01621459.1949.10483310

[44]   Richman, P.G. and Meister, A. (1975) Regulation of gamma-glutamyl-cysteine synthetase by nonallosteric feedback inhibition by glutathione. The Journal of Biological Chemistry, 250, 1422-1426.

 
 
Top