Serap DEMİREL, Mustafa USTA, Abdullah GÜLLER

Ekmeklik Buğday Bitkisinden Ribozom İnaktivite Eden Proteinin (Tritin) in silico Analizi

Ribozom inaktive eden proteinler (RIP’ler) ribozomal RNA’da spesifik bir adeninin depürünasyonundan sonra protein sentezini baskılyan enzimlerdir. Tritin RIP ailesinden RNA-N glikosidaz domainine sahip tip I RIP’lerden biridir. Mevcut çalışmada Kutluk-94 buğday çeşidinin yapraklarından tritini kodlayan cDNA izole edildi ve pGEM-T Easy vektöre klonlandı. Recombinant plazmid sekanslandı. Farklı biyoinformatik araçlar tritin proteininin özelliklerinin değerlendirilmesi için kullanıldı. Bazı monokotil bitkilerde toplamda 38 tritin benzeri sekans tespit edildi. Sonuçlar tritin proteininin diğer RIP’lerde bulunan RNA N-glikozidaz aktivitesi ile ilişkili korunmuş domaine (Ricin-A) sahip olduğunu ortaya koydu. Çoklu sekans hizalamaya analizi tritinin RNA N-glikozidaz aktivitesinde hayati rol oynayan korunmuş amino asitlere sahip olduğunu göstermiştir. Bizim çalışmamızda in silico analizlerden elde edilen sonuçlar tritin proteinin moleküler ve yapısal özellikleri hakkında diğer araştırmacılara bilgi sağlayacaktır.

Anahtar Kelimeler:

Ribozom inaktive eden protein, tritin, cDNA

In silico Analysis of Ribosome-Inactivating Protein (Tritin) from Common Wheat Plants (Triticum aestivum L.)

Ribosome-inactivating proteins (RIPs) are one of the enzymes that inhibit protein synthesis after depurination of a specific adenine in ribosomal RNA. The tritin is one of type I RIPs that include RNA-N glycosidase domain from RIP family. In the present study, cDNA encoding tritin from leaves of wheat Kutluk-94 cultivar was isolated and cloned into pGEM-T Easy vector. The recombinant plasmid was sequenced. The different bioinformatics tools were used for assessment of tritin protein characteristics. A total of 38 tritin-like sequences were identified in some monocot plants. Results showed that tritin protein have conserved domain (Ricin-A) found in other RIPs associated with RNA N-glycosidase activity and shows chancing homology to the RIPs in other plant species. According to multiple sequence alignment, tritin has conserved amino acids which are crucial role in RNA N-glycosidase activity. Our study illustrates that results obtained from in silico analyses could provide a perspective to another researcher about molecular and structural properties of tritin protein.

Keywords:

Ribosome-inactivating protein, tritin, cDNA,

PDF

___

Abbas, S. (2007). Cloning and expression of cDNA encoding ribosome inactivating proteins (Doctoral dissertation, UAS, Dharwad).
Ajji, P. K., Walder, K., & Puri, M. (2016). Functional analysis of a type-I ribosome inactivating protein balsamin from Momordica balsamina with anti-microbial and DNase activity. Plant foods for human nutrition, 71(3), 265-271.
Allahyari, H., Heidari, S., Ghamgosha, M., Saffarian, P., & Amani, J. (2017). Immunotoxin: A new tool for cancer therapy. Tumor Biology, 39(2), 1010428317692226.
Barbieri, L., Polito, L., Bolognesi, A., Ciani, M., Pelosi, E., Farini, V., ... & Stirpe, F. (2006). Ribosome-inactivating proteins in edible plants and purification and characterization of a new ribosome-inactivating protein from Cucurbita moschata. Biochimica et Biophysica Acta (BBA)-General Subjects, 1760(5), 783-792.
Benítez, J., Ferreras, J. M., Muñoz, R., Arias, Y., Iglesias, R., Córdoba-Díaz, M., ... & Girbés, T. (2005). Cytotoxicity of an ebulin l-anti-human CD105 immunotoxin on mouse fibroblasts (L929) and rat myoblasts (L6E9) cells expressing human CD105. Medicinal Chemistry, 1(1), 65-71.
Bertholdo-Vargas, L. R., Martins, J. N., Bordin, D., Salvador, M., Schafer, A. E., de Barros, N. M., ... & Carlini, C. R. (2009). Type 1 ribosome-inactivating proteins—Entomotoxic, oxidative and genotoxic action on Anticarsia gemmatalis (Hübner) and Spodoptera frugiperda (JE Smith)(Lepidoptera: Noctuidae). Journal of Insect Physiology, 55(1), 51-58.
Calixto, J. B. (2000). Efficacy, safety, quality control, marketing and regulatory guidelines for herbal medicines (phytotherapeutic agents). Brazilian Journal of medical and Biological research, 33(2), 179-189.
Chen, Z. C., White, R. F., Antoniw, J. F., & Lin, Q. (1991). Effect of pokeweed antiviral protein (PAP) on the infection of plant viruses. Plant Pathology, 40(4), 612-620. Choudhary, N., Kapoor, H. C., & Lodha, M. L. (2008). Cloning and expression of antiviral/ribosome-inactivating protein from Bougainvillea x buttiana. Journal of biosciences, 33(1), 91-101.
Dallal, J. A., & Irvin, J. D. (1978). Enzymatic inactivation of eukaryotic ribosomes by the pokeweed antiviral protein. FEBS letters, 89(2), 257-259. Virgilio, M. D., Lombardi, A., Caliandro, R., & Fabbrini, M. S. (2010). Ribosome-inactivating proteins: from plant defense to tumor attack. Toxins, 2(11), 2699-2737.
Domashevskiy, A. V., & Goss, D. J. (2015). Pokeweed antiviral protein, a ribosome inactivating protein: activity, inhibition and prospects. Toxins, 7(2), 274-298.
Donayre-Torres, A. J., Esquivel-Soto, E., Gutiérrez-Xicoténcatl, M. D., Esquivel-Guadarrama, F. R., & Gómez-Lim, M. A. (2009). Production and purification of immunologically active core protein p24 from HIV-1 fused to ricin toxin B subunit in E. coli. Virology Journal, 6(1), 1-11.
Duggar, B. M., & Armstrong, J. K. (1925). The effect of treating the virus of tobacco mosaic with the juices of various plants. Annals of the Missouri Botanical Garden, 12(4), 359-366.
Endo, Y., Mitsui, K., Motizuki, M., & Tsurugi, K. (1987). The mechanism of action of ricin and related toxic lectins on eukaryotic ribosomes. The site and the characteristics of the modification in 28 S ribosomal RNA caused by the toxins. Journal of Biological Chemistry, 262(12), 5908-5912.
Fabbrini, M. S., Katayama, M., Nakase, I., & Vago, R. (2017). Plant ribosome-inactivating proteins: Progesses, challenges and biotechnological applications (and a few digressions). Toxins, 9(10), 314.
Foissac, X., L. Savalle-Dumas, P. Gentit, M.J. Dulucq and T. Candresse. 2001. Polyvalent detection of fruit tree Tricho, Capillo and Faveaviruses by nested RT-PCR using degenerated and inosine containing primers (PDO RT-PCR). Acta Horticulturae, 357: 52-59.
Gasteiger, E., Hoogland, C., Gattiker, A., Wilkins, M. R., Appel, R. D., & Bairoch, A. (2005). Protein identification and analysis tools on the ExPASy server. The proteomics protocols handbook, 571-607.
Girbés, T., Ferreras, J. M., Arias, F. J., & Stirpe, F. (2004). Description, distribution, activity and phylogenetic relationship of ribosome-inactivating proteins in plants, fungi and bacteria. Mini reviews in medicinal chemistry, 4(5), 461-476.
Güller, A., Sipahioğlu, H. M., Usta, M., & Durak, E. D. (2018). Antiviral and Antifungal Activity of Biologically Active Recombinant Bouganin Protein from Bougainvillea spectabilis Willd. Journal of Agricultural Sciences, 24(2), 227-237.
Habuka, N., Akiyama, K., Tsuge, H., Miyano, M., Matsumoto, T., & Noma, M. (1990). Expression and secretion of Mirabilis antiviral protein in Escherichia coli and its inhibition of in vitro eukaryotic and prokaryotic protein synthesis. Journal of Biological Chemistry, 265(19), 10988-10992.
Habuka, N., Kataoka, J., Miyano, M., Tsuge, H., Ago, H., & Noma, M. (1993). Nucleotide sequence of a genomic gene encoding tritin, a ribosome-inactivating protein from Triticum aestivum. Plant molecular biology, 22(1), 171-176.
Hamshou, M., Shang, C., Smagghe, G., & Van Damme, E. J. (2016). Ribosome-inactivating proteins from apple have strong aphicidal activity in artificial diet and in planta. Crop Protection, 87, 19-24.
Hey, T. D., Hartley, M., & Walsh, T. A. (1995). Maize ribosome-inactivating protein (b-32)(homologs in related species, effects on maize ribosomes, and modulation of activity by pro-peptide deletions). Plant physiology, 107(4), 1323-1332.
Hogan, L. E., Vasquez, J., Hobbs, K. S., Hanhauser, E., Aguilar-Rodriguez, B., Hussien, R., ... & Henrich, T. J. (2018). Increased HIV-1 transcriptional activity and infectious burden in peripheral blood and gut-associated CD4+ T cells expressing CD30. PLoS pathogens, 14(2), e1006856.
Huang, M. X., Hou, P., Wei, Q., Xu, Y., & Chen, F. (2008). A ribosome-inactivating protein (curcin 2) induced from Jatropha curcas can reduce viral and fungal infection in transgenic tobacco. Plant Growth Regulation, 54(2), 115-123.
Iglesias, R., Citores, L., Ragucci, S., Russo, R., Di Maro, A., & Ferreras, J. M. (2016). Biological and antipathogenic activities of ribosome-inactivating proteins from Phytolacca dioica L. Biochimica et Biophysica Acta (BBA)-General Subjects, 1860(6), 1256-1264.
Kim, J. K., Jang, I. C., Wu, R., Zuo, W. N., Boston, R. S., Lee, Y. H., ... & Nahm, B. H. (2003). Co-expression of a modified maize ribosome-inactivating protein and a rice basic chitinase gene in transgenic rice plants confers enhanced resistance to sheath blight. Transgenic Research, 12(4), 475-484.
Krawetz, J. E., & Boston, R. S. (2000). Substrate specificity of a maize ribosome‐inactivating protein differs across diverse taxa. European Journal of Biochemistry, 267(7), 1966-1974.
Kumar, M. A., Timm, D. E., Neet, K. E., Owen, W. G., Peumans, W. J., & Rao, A. G. (1993). Characterization of the lectin from the bulbs of Eranthis hyemalis (winter aconite) as an inhibitor of protein synthesis. Journal of Biological Chemistry, 268(33), 25176-25183.
Lam, S. K., & Ng, T. B. (2001). First simultaneous isolation of a ribosome inactivating protein and an antifungal protein from a mushroom (Lyophyllum shimeji) together with evidence for synergism of their antifungal effects. Archives of Biochemistry and Biophysics, 393(2), 271-280.
Lam, S. K., & Ng, T. B. (2001b). Hypsin, a novel thermostable ribosome-inactivating protein with antifungal and antiproliferative activities from fruiting bodies of the edible mushroom Hypsizigus marmoreus. Biochemical and biophysical research communications, 285(4), 1071-1075.
Lapadula, W. J., & Ayub, M. J. (2017). Ribosome Inactivating Proteins from an evolutionary perspective. Toxicon, 136, 6-14.
Lapadula, W. J., Sanchez Puerta, M. V., & Juri Ayub, M. (2013). Revising the taxonomic distribution, origin and evolution of ribosome inactivating protein genes. PloS one, 8(9), e72825.
Liu, R. S., Yang, J. H., & Liu, W. Y. (2002). Isolation and enzymatic characterization of lamjapin, the first ribosome‐inactivating protein from cryptogamic algal plant (Laminaria japonica A). European journal of biochemistry, 269(19), 4746-4752.
Lombard, S., Helmy, M. E., & Pıéronı, G. (2001). Lipolytic activity of ricin from Ricinus sanguineus and Ricinus communis on neutral lipids. Biochemical Journal, 358(3), 773-781.
Madin, K., Sawasaki, T., Ogasawara, T., & Endo, Y. (2000). A highly efficient and robust cell-free protein synthesis system prepared from wheat embryos: plants apparently contain a suicide system directed at ribosomes. Proceedings of the National Academy of Sciences, 97(2), 559-564.
Mundy, J., Leah, R., Boston, R., Endo, Y., & Stirpe, F. (1994). Genes encoding ribosome-inactivating proteins. Plant Molecular Biology Reporter, 12(2), S60-S62.
Nielsen, K., & Boston, R. S. (2001). Ribosome-inactivating proteins: a plant perspective. Annual review of plant biology, 52(1), 785-816.
Olsnes, S., & Pihl, A. (1973a). Different biological properties of the two constituent peptide chains of ricin a toxic protein inhibiting protein synthesis. Biochemistry, 12(16), 3121-3126.
Olsnes, S., & Pihl, A. (1973b). Isolation and Properties of Abrin: a Toxic Protein Inhibiting Protein Synthesis: Evidence for Different Biological Functions of Its Two Constituent‐Peptide Chains. European journal of biochemistry, 35(1), 179-185.
Parkash, A., Ng, T. B., & Tso, W. W. (2002). Isolation and characterization of luffacylin, a ribosome inactivating peptide with anti-fungal activity from sponge gourd (Luffa cylindrica) seeds. Peptides, 23(6), 1019-1024.
Peumans, W. J., Shang, C., & Van Damme, E. J. (2014). Updated model of the molecular evolution of RIP genes. Ribosome‐inactivating Proteins: Ricin and Related Proteins, 134-150.
Peumans, W. J., Hao, Q., & Van Damme, E. J. (2001). Ribosome‐inactivating proteins from plants: more than RNA N‐glycosidases?. The FASEB Journal, 15(9), 1493-1506.
Peumans, W. J., & Van Damme, E. J. (2010). Evolution of plant ribosome-inactivating proteins. In Toxic plant proteins (pp. 1-26). Springer, Berlin, Heidelberg.
Praveen, S., Tripathi, S., & Varma, A. (2001). Isolation and characterization of an inducer protein (Crip-31) from Clerodendrum inerme leaves responsible for induction of systemic resistance against viruses. Plant Science, 161(3), 453-459.
Ruggiero, A., Chambery, A., Di Maro, A., Mastroianni, A., Parente, A., & Berisio, R. (2007). Crystallization and preliminary X-ray diffraction analysis of PD-L1, a highly glycosylated ribosome inactivating protein with DNase activity. Protein and peptide letters, 14(4), 407-709.
Rumiyati, N. A. W., Sismindari-Lukitaningsih, E., & Yuliati, T. (2014). Potential of ribosome-inactivating proteins (RIPs) of Mirabilis jalapa L. as an antiacne: effect on proliferation of cultured sebocyte cells and its antibacterial activities against Propionibacterium acnes and Staphylococcus epidermidis. International Journal of Pharmaceutical Chemistry, 4, 130-133.
Shahidi-Noghabi, S., Van Damme, E. J., & Smagghe, G. (2008). Carbohydrate-binding activity of the type-2 ribosome-inactivating protein SNA-I from elderberry (Sambucus nigra) is a determining factor for its insecticidal activity. Phytochemistry, 69(17), 2972-2978.
Shang, C., Rougé, P., & Van Damme, E. J. (2016). Ribosome inactivating proteins from Rosaceae. Molecules, 21(8), 1105.
Sharma, N., Park, S. W., Vepachedu, R., Barbieri, L., Ciani, M., Stirpe, F., ... & Vivanco, J. M. (2004). Isolation and characterization of an RIP (ribosome-inactivating protein)-like protein from tobacco with dual enzymatic activity. Plant Physiology, 134(1), 171-181.
Shih, N. R., McDonald, K. A., Jackman, A. P., Girbés, T., & Iglesias, R. (1997). Bifunctional plant defence enzymes with chitinase and ribosome inactivating activities from Trichosanthes kirilowii cell cultures. Plant Science, 130(2), 145-150.
Shu, S. H., Xie, G. Z., Guo, X. L., & Wang, M. (2009). Purification and characterization of a novel ribosome-inactivating protein from seeds of Trichosanthes kirilowii Maxim. Protein expression and purification, 67(2), 120-125.
Sipahioğlu, H. M., Kaya, ˙I., Usta, M., Ünal, M., Özcan, D., Özer, M., Güller, A., and Pallas, V., (2017). Pokeweed (Phytolacca americana L.) antiviral protein inhibits Zucchini yellow mosaic virus infection in a dose-dependent manner in squash plants. Turkish Journal of Agriculture and Forestry, 41, 256–262.
Song, S. K., Choi, Y., Moon, Y. H., Kim, S. G., Do Choi, Y., & Lee, J. S. (2000). Systemic induction of a Phytolacca insularis antiviral protein gene by mechanical wounding, jasmonic acid, and abscisic acid. Plant molecular biology, 43(4), 439-450.
Stirpe, F. (2004). Ribosome-inactivating proteins. Toxicon, 44(4), 371-383.
Stirpe, F., & Battelli, M. G. (2006). Ribosome-inactivating proteins: progress and problems. Cellular and Molecular Life Sciences CMLS, 63(16), 1850-1866.
Vivanco, J. M., Savary, B. J., & Flores, H. E. (1999). Characterization of two novel type I ribosome-inactivating proteins from the storage roots of the Andean crop Mirabilis expansa. Plant Physiology, 119(4), 1447-1456.
Wang, S., Zhang, Y., Liu, H., He, Y., Yan, J., Wu, Z., & Ding, Y. (2012). Molecular cloning and functional analysis of a recombinant ribosome-inactivating protein (alpha-momorcharin) from Momordica charantia. Applied microbiology and biotechnology, 96(4), 939-950.
Wei, G. Q., Liu, R. S., Wang, Q., & Liu, W. Y. (2004). Toxicity of two type II ribosome‐inactivating proteins (cinnamomin and ricin) to domestic silkworm larvae. Archives of Insect Biochemistry and Physiology: Published in Collaboration with the Entomological Society of America, 57(4), 160-165.
Yao, Q. Z., Yu, M. M., Ooi, L. S., Ng, T. B., Chang, S. T., Sun, S. S., & Ooi, V. E. (1998). Isolation and characterization of a type 1 ribosome-inactivating protein from fruiting bodies of the edible mushroom (Volvariella volvacea). Journal of agricultural and food chemistry, 46(2), 788-792.
Zhu, F., Xu, M., Wang, S., Jia, S., Zhang, P., Lin, H., & Xi, D. (2012). Prokaryotic expression of pathogenesis related protein 1 gene from Nicotiana benthamiana: antifungal activity and preparation of its polyclonal antibody. Biotechnology letters, 34(5), 919-924.
Zhu, F., Yuan, S., Zhang, Z. W., Qian, K., Feng, J. G., & Yang, Y. Z. (2016). Pokeweed antiviral protein (PAP) increases plant systemic resistance to Tobacco mosaic virus infection in Nicotiana benthamiana. European journal of plant pathology, 146(3), 541-549.
Zhu, F., Zhang, P., Meng, Y. F., Xu, F., Zhang, D. W., Cheng, J., ... & Xi, D. H. (2013). Alpha-momorcharin, a RIP produced by bitter melon, enhances defense response in tobacco plants against diverse plant viruses and shows antifungal activity in vitro. Planta, 237(1), 77-88.
Zhu, F., Zhou, Y. K., Ji, Z. L., & Chen, X. R. (2018). The plant ribosome-inactivating proteins play important roles in defense against pathogens and insect pest attacks. Frontiers in Plant Science, 9, 146.