Cytotoxic effects of an acidic Ribosome-inactivating Protein like protein isolated from Mirabilis jalapa L. leaves on cancer cell-lines
Most purification approaches of Ribosome-inactivating Proteins (RIPs) used a cation exchange chromatography resulting highly basic proteins, such as MJ-30 isolated from Mirabilis jalapa leaves. The M.jalapa unbounded protein fractions of the column showed cleavage activity on supercoiled double stranded DNA and possesed cytotoxic effects on cancer cell lines. So the purification and characterization of the acidic protein was needed to be done.
Total proteins of M.jalapa leaves was collected from precipitating process of the extract using ammonium sulphate untill saturated. After dialysis, the proteins was applied into the weak base anion exchanger Ionenaustaucher Typ II. The bounded proteins was eluted with 0.0 to 0.5 M NaCl gradient in phosphate buffer. Active protein fractions were determined by supercoilded DNA cleavage activity and then cytotoxic activity was testes.
Results showed that protein fraction eluted at 0.35 – 0.40 M NaCl possesed cleavage activity on supercoiled DNA. The acidic protein, MJ-C, was toxic on HeLa, Myeloma and T47D cell lines, with the LC50 of 14.3 μg/mL, 7.4 μg/mL and 27.8 μg/mL, respectively. The MJ-C was more potent than the basic protein MJ-30.
Key words: acidic RIP, cytotoxic, Mirabilis jalapa
Barbieri, L., Batteli, M. G., and Stirpe, F., 1993, Ribosome-inactivating proteins from plants, Biochem. Biophys. Acta., 1154: 237-282.
Battelli, M. G., 2004, Cytotoxicity and toxicity to animals and humans of ribosome-inactivating proteins, Mini-review, Med Chem., 4: 513-521.
Berenson, J. R., and Miguel, J. S., 2003, Multiple myeloma, htpp//www.multiplemyeloma.com, Nopember 2005.
Bolognesi, A., Polito, L., Lubelli, C., Barbieri, L., Parante, A., dan Stirpe, F., 2002, Ribosomeinactivating and Adenine Polynucleotide Glycosylase Activities in Mirabilis jalapa L Tissues, J. Biol. Chem., 277: 13709 - 716.
Desaintes, C., Goyat, S., Garbay, S., Yaniv, M., and Thierry, F., 1999, Papillomavirus E2 induces p53-independent apoptosis in HeLa cell, Oncogene, 18: 4533-4545
Hartley, M. R., Chaddock, J. A., and Bonnes, M. R., 1996, The structure and Fuction of ribosomeinactivating proteins, Trends Plant Sci Rev., 1(8): 254 - 258
Ikawati, Z., Sudjadi, Elly, W., Pusputasari, D., and Sismindari, 2003, Induction of apoptosis by protein fraction isolated from the leaves of Mirabilis jalapa L on HeLa and Raji cell line, OPEM, 3 (3): 151-156.
Ikawati, Z., Sudjadi and Sismindari, 2006, Cytotoxic against tumor cell lines of a Ribosomeinactivating like protein isolated from leaves of Mirabilis jalapa L , Malaysian J Pharm.Sciences, 4 (1) in press.
Layne, E., 1957, Spectrophotometric and turbidimetric methods for measuring proteins, in Method Enzymol., 3: 477, Colowick and Kaplan, Academic Press, New York.
LingJ., Lui, W., and Wang, T. P., 1994, Cleavage of supercoiled double stranded DNA by several Ribosome-inactivating Proteins in vitro, FEBS Letters, 345: 143- 146.
Mundy, J., Leah, R., Boston,R., Endo, Y., and Stirpe, F., 1994, Genes encoding ribosomeinactivating proteins, Plant Mol. Biol. Rep., 12: 60-62.
Narayanan, S., Surolia, A., and Karande, A., 2004, Ribosome-inactivating proteins and apoptosis: abrin causes cell death via mitochondrial pathway in Jurkat cells, Biochem. J., 377: 233-240.
Parkash, A., Ng, T. B., and Tso, W. W., 2002, Purification and characterization of charantin, a napin-like ribosome-inactivating peptide from bitter gourd (Momordica charantia) seeds, J.Peptide Res, 59: 197-202.
Peumans, W., Hao, Q., and van Damme, E., 2001, Ribosome-inactivating proteins from plants: more than RNA N-glycosidase, J FASEB, 15: 1493-1506.
Sismindari and Lord, J. M., 2000, Ribosome-inactivating proteins RNA N-glycosidase activity of Mirabilis jalapa L., Morinda citrifolia L., and Carica papaya L., Indon. J. Biotech., 342-345.
Stirpe, F., Gasperi-Campani, A., Barbieri, L., Falasca, A., Abbondanza, A., Stevens, W. A., 1983, Ribosome-inactivating Proteins from the Seeds of Saponaria officinalis L. (soapwort), of Argositemma githago L (corn cocle) and Asparagus officinalis L. (asparagus) and from the Latex of Hura crepitans L. (sandbox tree)., Biochem. J., 216, 617 - 625
Stirpe, F., Barbieri, R., Batteli, M. G., Soria, M., and Lappi, D. A., 1992, RIP from plants: Present status and future prospect, Review, Biotech., 10: 105-109.
Sudjadi, Sismindari, Herawati, T., Prasetyowati, A. T., 2003, Pemurnian Ribosome-inactivating Proteein (RIP) dari daun Mirabilis jalapa L dengan kolom CM-Sepharose CL-6B dan Sephacryl S-300HR, MFI, 14 (2): 316-321.
Sudjadi dan Ikawati, Z., 2003, Efek sitotoksik protein 30 kD dari daun Mirabilis jalapa L pada kultur sel HeLa, myeloma, SiHa, Vero dan mononuklear darah perifer dan uji stabilitasnya, Laporan Penelitian Project-Grand Que Project Fakultas Farmasi UGM.
Takanami, Y., Kuwata, S., Ikeda, T., and Kubo, S., 1990, Purification and characterization of the anti-plant viral protein from Mirabilis jalapa L., Ann. Phytopathol. Soc. Jpn., 56: 488-494.
Wosniak, M. A., and Keely, P. J., 2005, Use of tree-dimensional collagen gels to study mechanotransduction in T47D breast epithelial cells, Biol. Proced Online, 7: 144-146. d
- There are currently no refbacks.
Copyright (c) 2017 INDONESIAN JOURNAL OF PHARMACY
This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.
Indonesian J Pharm indexed by:View My Stats