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Korean Society for Biochemistry and Molecular Biology (생화학분자생물학회)
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Tat-mediated Protein Transduction of Human Brain Pyridoxine-5-P Oxidase into PC12 Cells

  • Kim, So-Young (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • An, Jae-Jin (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Kim, Dae-Won (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Choi, Soo-Hyun (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Lee, Sun-Hwa (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Hwang, Seok-Il (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Kwon, Oh-Shin (Department of Biochemistry, Kyungpook National University) ;
  • Kang, Tae-Cheon (Department of Anatomy, College of Medicine, Hallym University) ;
  • Won, Moo-Ho (Department of Anatomy, College of Medicine, Hallym University) ;
  • Cho, Sung-Woo (Department of Biochemisuy and Molecular Biology, University of Ulsan College of Medicine) ;
  • Park, Jin-Seu (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Eum, Won-Sik (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Lee, Kil-Soo (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University) ;
  • Choi, Soo-Young (Department of Biomedical Sciences and Research Institute for Bioscience and Biotechnology, Hallym University)
  • Received : 2005.10.04
  • Accepted : 2005.11.02
  • Published : 2006.01.31
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Abstract

Pyridoxine-5-P oxidase catalyses the terminal step in the biosynthesis of pyridoxal-S-P, the biologically active form of vitamin $B_6$ Which acts as an essential cofactor. Here, a human brain pyridoxine-5-P oxidase gene was fused with a gene fragment encoding the HIV-1 Tat protein transduction domain (RKKRRQRRR) in a bacterial expression vector to produce a genetic in-frame Tat-pyridoxine-5-P oxidase fusion protein. Expressed and purified Tat-pyridoxine-5-P oxidase fusion protein transduced efficiently into PC12 cells in a time- and dose-dependent manner when added exogenously to culture media. Once inside the cells, the transduced Tat-pyridoxine-5-P oxidase protein showed catalytic activity and was stable for 48 h. Moreover, the formation of pyridoxal-5-P was increased by adding exogenous Tat-pyridoxine-5-P oxidase to media pre-treated with the vitamin $B_6$ precursor pyridoxine. In addition, the intracellular concentration of pyridoxal-S-P was markedly increased when Tat-pyridoxal kinase was transduced together with Tat-pyridoxine-5-P oxidase into cells. These results suggest that the transduction of Tat-pyridoxine-5-P oxidase fusion protein presents a means of regulating the level of pyridoxal-5-P and of replenishing this enzyme in various neurological disorders related to vitamin $B_6$.

Keywords

References

  1. Allgood, V. E., Oakley, R. H. and Cidlowski, J. A. (1993) Modulation by vitamin $B_6$ of glucocorticoid receptor-mediated gene expression requires transcription factors in addition to the glucocorticoid receptor. J. Biol. Chem. 268, 20870-20876.
  2. Bahn, J. H., Kwon, O. S., Joo, H. M., Jang, S. H., Park, J., Hwang, I. K., Kang, T. C., Won, M. H., Kwon, H. Y., Kwok, F., Kim, H. B., Cho, S. W. and Choi, S. Y. (2002) Immunohistochemical studies of brain pyridoxine-5-phosphate oxidase. Brain Res. 925, 159-168. https://doi.org/10.1016/S0006-8993(01)03271-1
  3. Black, A. L., Guirard, B. M. and Snell, E. E. (1977) Increased muscle phosphorylase in rats fed high levels of vitamin $B_6$. J. Nutr. 107, 1962-1968. https://doi.org/10.1093/jn/107.11.1962
  4. Bradford, M. A. (1976) A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248-254. https://doi.org/10.1016/0003-2697(76)90527-3
  5. Brewis, N., Phelan, A., Webb, J., Drew, J., Elliott, G. and O'Hare, P. (2000) Evaluation of VP22 spread in tissue culture. J. Viol. 74, 1051-1056.
  6. Cashman, S. M., Morris, D. J. and Kumar-Singh, R. (2003) Evidence of protein transduction but not intracellular transport by protein fused to HIV Tat in retinal cell culture and in vivo. Mol. Ther. 8, 130-142. https://doi.org/10.1016/S1525-0016(03)00131-X
  7. Choi, J. D., Bowers-Komro, M., Davis, D. M., Edmondson, D. E. and McCormick, D. B. (1983) Kinetic properties of pyridoxamine (pyridoxine)-5'-phosphate oxidase from rabbit liver. J. Biol. Chem. 258, 840-845.
  8. Choi, S. Y., Churchich, J. E., Zaiden, E. and Kwok, F. (1987) Brain pyridoxine-5-P oxidase. Modulation of its catalytic activity by reaction with pyridoxal-5-P analogs. J. Biol. Chem. 262, 12013-12017.
  9. Choi, S. Y., Lee, S. J. and Cho, S. W. (1992) Catalytic and structure properties of brain pyridoxal kinase. J. Biochem. Mol. Biol. 25, 624-630.
  10. Churchich, J. E. (1984) Brain pyridoxine-5-phosphate oxidase. A dimeric enzyme containing one FMN site. Eur. J. Biochem. 138, 327-332. https://doi.org/10.1111/j.1432-1033.1984.tb07918.x
  11. Dakshinamurti, K., Paulose, C. S., Viswanathan, M., Siow, Y. L., Sharma, S. K. and Bolster, B. (1990) Neurobiology of pyridoxine. Ann. N. Y. Acad. Sci. 585, 128-144. https://doi.org/10.1111/j.1749-6632.1990.tb28049.x
  12. Derrosi, D., Joliot, A. H., Chassaing, G. and Prochiantz, A. (1994) The third helix of the Antennapedia homeodomain translocates through biological membranes. J. Biol. Chem. 269, 10444-10450.
  13. Elliott, G. and O'Hare, P. (1997) Intercellular trafficking and protein delivery by a herpes virus structural protein. Cell 88, 223-233. https://doi.org/10.1016/S0092-8674(00)81843-7
  14. Eum, W. S., Choung, I. S., Kim, A. Y., Lee, Y. J., Kang, J. H., Park, J., Lee, K. S., Kwon, H. Y. and Choi, S. Y. (2002) Transduction efficacy of Tat-Cu,Zn-superoxide dismutase is enhanced by copper ion recovery of the fusion protein. Mol. Cells 13, 334-340.
  15. Eum, W. S., Choung, I. S., Li, M. Z., Kang, J. H., Kim, D.W., Park, J., Kwon, H. Y. and Choi, S. Y. (2004a) HIV-1 Tat mediated protein transduction of Cu,Zn-superoxide dismutase into pancreatic ${\beta}$ cells in vitro and in vivo. Free Radic. Biol. Med. 37, 339-349. https://doi.org/10.1016/j.freeradbiomed.2004.04.036
  16. Eum, W. S., Kim, D. W., Hwang, I. K., Yoo, K. Y., Kang, T. C., Jang, S. H., Choi, H. S., Choi, S. H., Kim, Y. H., Kim, S. Y., Kwon, H. Y., Kang, J. H., Kwon, O. S., Cho, S. W., Lee, K. S., Park, J., Won, M. H. and Choi, S.Y. (2004b) In vivo protein transduction: biologically active intact PEP-1- superoxide dismutase fusion protein efficiently protects against ischemic insults. Free Radic. Biol. Med. 37, 1656-1669. https://doi.org/10.1016/j.freeradbiomed.2004.07.028
  17. Frankel, A. D. and Pabo, C. O. (1988) Cellular uptake of the tat protein from human immunodeficiency virus. Cell 55, 1189-1193. https://doi.org/10.1016/0092-8674(88)90263-2
  18. Glenn, G. M., Krober, M. S., Kelly, P., McCarty, J. and Weir, M. (1995) Pyridoxine as therapy in theophylline-induced seizures. Ver. Hum. Nerol. 395, 177-194.
  19. Gospe, S. M. Jr., Olin, K. L. and Keen, C. L. (1994) Reduced GABA synthesis in pyridoxine dependent seizures. Lancet 343, 1133-1134. https://doi.org/10.1016/S0140-6736(94)90236-4
  20. Han, K. H., Jeon, M. J., Kim, K. A., Park, J. and Choi, S. Y. (2000) Efficient intracellular delivery of GFP by homeodomains of Drosophila fushi-tarazu and engrailed proteins. Mol. Cells 10, 727-731.
  21. Han, K. H., Jeon, M. J., Kim, S. H., Ki, D., Bahn, J. H., Lee, K. S., Park, J. and Choi, S. Y. (2001) Efficient intracellular delivery of an exogenous protein GFP with genetically fused basic oligopeptides. Mol. Cells 12, 267-271.
  22. Jin, L. H., Bahn, J. H., Eum, W. S., Kwon, H. Y., Jang, S. H., Han, K. H., Kang, T. C., Won, M. H., Kang, J. H., Cho, S. W., Park, J. and Choi, S. Y. (2001) Transduction of human catalase mediated by an HIV-1 Tat protein basic domain and argininerich peptides into mammalian cells. Free Radic. Biol. Med. 31, 1509-1519. https://doi.org/10.1016/S0891-5849(01)00734-1
  23. Kabouridis, P. S. (2003) Biological applications of protein transduction technology. TRENDS Biotechnology 21, 498-503. https://doi.org/10.1016/j.tibtech.2003.09.008
  24. Kang, H. T., Bang, W. K. and Yu, Y. G. (2004) Identification and characterization of a novel angiostatin-binding protein by the display cloning method. J. Biochem. Mol. Biol. 37, 159-166l. https://doi.org/10.5483/BMBRep.2004.37.2.159
  25. Kang, T. C., Park, S. K., Hwang, I. K., An, S. J., Bahn, J. H., Kim, A. Y., Choi, S.Y., Kwon, O. S., Baek, N. I., Lee, H. Y. and Won, M. H. (2002a) Chronological changes in pyridoxine- 5'-phosphate oxidase immunoreactivity in the seizure-sensitive gerbil hippocampus. J. Neurosci. Res. 68, 785-791. https://doi.org/10.1002/jnr.10280
  26. Kang, T. C., Park, S. K., Hwang, I. K., An, S. J., Bahn, J. H., Kim, D. W., Choi, S. Y., Kwon, O. S., Baek, N. I., Lee, H. Y. and Won, M. H. (2002b) Changes in pyridoxal kinase immunoreactivity in the gerbil hippocampus following spontaneous seizure. Brain Res. 957, 242-250. https://doi.org/10.1016/S0006-8993(02)03566-7
  27. Kazarinoff, M. N. and McCormick, D. B. (1975) Rabbit liver pyridoxamine (pyridoxine) 5'-phosphate oxidase. Purification and properties. J. Biol. Chem. 250, 3436-3442.
  28. Kim, D. W., Eum, W. S., Jang, S. H., Kim, S. Y., Choi, H. S., Choi, S. H., An, J. J., Lee, S. H., Lee, K. S., Han, K., Kang, T. C., Won, M. H., Kang, J. H., Kwon, O. S., Cho, S. W., Kim, T. Y., Park, J. and Choi, S. Y. (2005a) Transduced Tat- SOD fusion protein protects against ischemic brain injury. Mol. Cells 19, 88-96.
  29. Kim, D. W., Kim, C. K., Choi, S. H., Choi, H. S., Kim, S. Y., An, J. J., Lee, S. R., Lee, S. H., Kwon, O. S., Kang, T. C., Won, M. H., Cho, Y. J., Cho, S. W., Kang, J. H., Kim, T. Y., Lee, K. S., Park, J., Eum, W. S. and Choi, S. Y. (2005b) Tatmediated protein transduction of human brain pyridoxal kinase into PC12 cells. Biochimie 87, 481-487. https://doi.org/10.1016/j.biochi.2004.12.004
  30. Kim, H. J., Yang, S. J., Kim, Y. S. and Kim, T. U. (2003) Cobalt chloride-induced apoptosis and extracellular signal-regulated protein kinase activation in human cervical cancer HeLa cells. J. Biochem. Mol. Biol. 36, 468-474. https://doi.org/10.5483/BMBRep.2003.36.5.468
  31. Kwok, F. and Churchich, J. E. (1979) Brain pyridoxal kinase: purification, substrate specificities, and sensitized photodestruction of an essential histidine. J. Biol. Chem. 254, 6489-6495.
  32. Kwok, F. and Churchich, J. E. (1980) Interaction between pyridoxal kinase and pyridoxine-5-P oxidase, two enzymes involved in the metabolism of vitamin $B_6$. J. Biol. Chem. 255, 882-887.
  33. Kwon, H. Y., Eum, W. S., Jang, H. W., Kang, J. H., Ryu, J., Lee, B. R., Jin, L. H., Park, J. and Choi, S. Y. (2000) Transduction of Cu,Zn-superoxide dismutase mediated by an HIV-1 Tat protein basic domain into mammalian cells. FEBS Lett. 485, 163-167. https://doi.org/10.1016/S0014-5793(00)02215-8
  34. Kwon, O. S., Kwok, F. and Churchich, J. E. (1991) Catalytic and regulatory properties of native and chymotrypsin-treated pyridoxine-5-phosphate oxidase. J. Biol. Chem. 266, 22136- 22140.
  35. Lam, H. M. and Winkler, M. E. (1992) Characterization of the complex pdxH-tyrS operon of E. coli K-12 and pleiotropic phenotypes caused by pdxH insertion mutations. J. Bacteriol. 174, 6033-6045. https://doi.org/10.1128/jb.174.19.6033-6045.1992
  36. McCormick, D. B., Gregory, M. E. and Snell, E. E. (1961) Pyridoxal phosphokinase. I. Assay distribution, purification, and properties. J. Biol. Chem. 236, 2076-2084.
  37. Meisler, N. T. and Thanassi, J. W. (1980) Pyridoxine kinase, pyridoxine phosphate phosphatase and pyridoxine phosphate oxidase activities in control and B-6-deficient rat liver and brain. J. Nutr. 110, 1965-1975. https://doi.org/10.1093/jn/110.10.1965
  38. Park, J., Ryu, J., Kim, K. A., Lee, H. J., Bahn, J. H., Han, K., Choi, E. Y., Lee, K. S., Kwon, H. Y. and Choi, S. Y. (2002a) Mutational analysis of a human immunodeficiency virus type 1 Tat protein transduction domain which is required for delivery of an exogenous protein into mammalian cells. J. Gen. Virol. 83, 1173-1181. https://doi.org/10.1099/0022-1317-83-5-1173
  39. Park, J., Ryu, J., Jin, L. H., Bahn, J. H., Kim, J. A., Yoon, C. S., Kim, D. W., Han, K. H., Eum, W. S., Kwon, H. Y., Kang, T. C., Won, M. H., Kang, J. H., Cho, S. W. and Choi, S. Y. (2002b) 9-Polylysine protein transduction domain: Enhanced penetration efficiency of superoxide dismutase into mammalian cells and skin. Mol. Cells 13, 202-208.
  40. Richard, J. P., Melikov, K., Vives, E., Ramos, C., Verbeure, B., Gait, M. J., Chernomordik, L. V. and Lebleu, B. (2003) Cellpenetrating peptides: A reevaluation of the mechanism of cellular uptake. J. Biol. Chem. 278, 585-590. https://doi.org/10.1074/jbc.M209548200
  41. Schwartze, S. R., Ho, A., Vocero-Akbani, A. and Dowdy, S. F. (1999) In vivo protein transduction: delivery of a biologically active protein into the mouse. Science 285, 1569-1572. https://doi.org/10.1126/science.285.5433.1569
  42. Schwartze, S. R., Hruska, K. A. and Dowdy, S. F. (2000) Protein transduction: unrestricted delivery into all cell? Trends in Cell Biol. 10, 290-295. https://doi.org/10.1016/S0962-8924(00)01771-2
  43. Snell, E. E. (1990) Vitamin $B_6$ and decarboxylation of histidine. Ann. N. Y. Acad. Sci. 585, 1-12. https://doi.org/10.1111/j.1749-6632.1990.tb28037.x
  44. Wadia, J. S. and Dowdy, S. (2002) Protein transduction technology. Curr. Opin. Biotechnol. 13, 52-56. https://doi.org/10.1016/S0958-1669(02)00284-7
  45. Waymire, K. G., Mahuren, J. D., Jaje, J. M., Guilarte, T. R., Coburn, S. P. and MacGregor, G. R. (1995) Mice lacking tissues non-specific alkaline phosphatase die from seizures due to defective metabolism of vitamin $B_6$. Nat. Genet. 11, 45-51. https://doi.org/10.1038/ng0995-45
  46. Zhao, G. and Winkler, M. E. (1995) Kinetic limitation and cellular amount of pyridoxamine (pyridoxine) 5'-phosphate oxidase of E. coli K-12. J. Bacteriol. 177, 883-891. https://doi.org/10.1128/jb.177.4.883-891.1995

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