PENGUJIAN POTENSI ALERGENITAS COAT PROTEIN OF SUGARCANE MOZAIC VIRUS PADA TANAMAN TEBU TRANSGENIK
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Abstract
Sugarcane resistant to sugarcane mosaic virus (SCMV) was developed by overexpression of gene for coat protein (CP). Therefore, this study aimed at investigating the potential allergenicity of CP-SCMV in transgenic sugarcane. Allergenicity was assessed by analysis in silico and in vitro. In silico analysis using AllergenOnline FASTA alignment of full-length CP-SCMV amino acid showed that the protein had no similarity with allergen protein. However, the alignment using 80 mer CP-SCMV showed over 35% similarity, but this result was considered as false positive. In silico analysis on digestion capability of protease found the cutting sites of CP-SCMV by pepsin, trypsin and chymotrypsin. This result was further confirmed by in vitro gastrointestinal digestion in that CP-SCMV was digested by pepsin and trypsin. Although CP-SCMV was less degraded by in vitro heat treatment and quantitatively underwent slight decrease after 30-minute heating on 90 ºC, the protein might lose its function. These results indicated that CP-SCMV was considered having no potential allergen in transgenic sugarcane resistant to SCMV.
Tebu tahan sugarcane mosaic virus (SCMV) dirakit melalui overekspresi gen untuk coat protein (CP). Oleh karena itu, penelitian ini bertujuan menguji alergenitas CP-SCMV pada tebu transgenik. Pengujian alergenitas dilakukan melalui analisis in silico dan in vitro. Hasil analisis in silico dengan pensejajaran AllergenOnline FASTA full-length asam amino CP-SCMV menunjukkan tidak ada kesamaan dengan protein alergen. Namun demikian pada pensejajaran 80 mer, CP-SCMV mempunyai kemiripan di atas 35% dengan alergen, tetapi hasil ini memiliki kecenderungan positif palsu. Analisis in silico terhadap kemampuan cerna protease ditemukan adanya sisi pemotongan CP-SCMV oleh enzim pensin, trypsin dan chymotrypsin. Hasil ini dikonfirmasi lebih lanjut dengan analisis in vitro pencernaan gastrointestinal yang menunjukkan bahwa CP-SCMV terdegradasi oleh pepsin dan trypsin. Walaupun hasil analisis in vitro menunjukkan CP-SCMV kurang dipengaruhi oleh perlakuan panas dan hanya sedikit berkurang pada pemanasan 90 ºC selama 30 menit, tetapi mungkin fungsi protein telah rusak. Hasil penelitian ini menyimpulkan bahwa CP-SCMV pada tanaman tebu transgenik tahan virus tidak berpotensi sebagai alergen.
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References
A alberse RC (2000) Structural biology of allergens. J Allergy Clin Immunol 106: 228–238. doi: 10.1067/mai.2000.108434
Addy HS, Nurmalasari, Wahyudi AHS, Sholeh A, Anugrah C, Iriyanto FES, Darmanto W, Sugiharto B (2017) Detection and response of sugarcane against the infection of sugarcane mosaic virus (SCMV) in Indonesia. Agronomy 7: 50. doi: 10.3390/agronomy7030050
Ahn J, Cao MJ, Yu YQ, Engen JR (2013) Accessing the reproducibility and specificity of pepsin and other aspartic proteases. Biochim Biophys Acta 1834: 1222–1229. doi: 10.1016/j.bbapap.2012.10.003
Alonso MG (2013) Safety assessment of food and feed derived from GM crops: using problem formulation to ensure “fit for purpose” risk assessments. Collection of Biosafety Reviews 8: 72–101. Corpus ID: 201851937
Apriasti R, Widyaningrum S, Hidayati WN, Sawitri WD, Darsono N, Hase T, Sugiharto B (2018) Full sequence of the coat protein gene is required for the induction of pathogen-derived resistance against sugarcane mosaic virus in transgenic sugarcane. Mol Biol Rep 45: 2749–2758. doi: 10.1007/s11033-018-4326-1
Braidwood L, Müller SY, Baulcombe D (2019) Extensive recombination challenges the utility of sugarcane mosaic virus phylogeny and strain typing. Sci Rep 9: 20067. doi: 10.1038/s41598-019-56227-y
Cressman RF, Ladics G (2009) Further evaluation of the utility of “Sliding Window” FASTA in predicting cross-reactivity with allergenic proteins. Regul Toxicol Pharmacol 54: S20–S25. doi: 10.1016/j.yrtph.2008.11.006
Darsono N, Azizah NN, Putranty KM, Astuti NT, Addy HS, Darmanto W, Sugiharto B (2018) Production of a polyclonal antibody against the recombinant coat protein of the sugarcane mosaic virus and its application in the immunodiagnostic of sugarcane. Agronomy 8: 93. doi: 10.3390/agronomy8060093
Farias DF, Viana MP, Oliveira GR, Santos VO, Pinto CEM, Viana DA, Vasconcelos IM, Grossi-de-Sa MF, Carvalho AFU (2015) Food safety assessment of Cry8Ka5 mutant protein using Cry1Ac as a control Bt protein. Food Chem Toxicol 81: 81–91. doi: 10.1016/j.fct.2015.04.008
Fu Z, Akula S, Thorpe M, Hellman L (2021) Marked difference in efficiency of the digestive enzymes pepsin, trypsin, chymotrypsin, and pancreatic elastase to cleave tightly folded proteins. Biol Chem 402: 861–867. doi: 10.1515/hsz-2020-0386
Gan J, Chen H, Liu J, Wang Y, Nirasawa S, Cheng Y (2016) Interactions of ?-conglycinin (7S) with different phenolic acids—impact on structural characteristics and proteolytic degradation of proteins. Int J Mol Sci 17: 1671. doi: 10.3390/ijms17101671
Giraldo PA, Shinozuka H, Spangenberg GC, Cogan NOI, Smith KF (2019) Safety assessment of genetically modified feed: Is there any difference from food? Front Plant Sci 10: 1592. doi: 10.3389/fpls.2019.01592
Goodman RE, Tetteh AO (2011) Suggested improvements for the allergenicity assessment of genetically modified plants used in foods. Curr Allergy Asthma Rep 11: 317–324. doi: 10.1007/s11882-011-0195-6
Goodman RE (2014) Biosafety: evaluation and regulation of genetically modified (GM) crops in the United States. J Huazhong Agricultural University 33: 85–114
Hidayati WN, Apriasti R, Addy HS, Sugiharto B (2021) Distinguishing resistances of transgenic sugarcane generated from RNA interference and pathogen?derived resistance approaches to combating sugarcane mosaic virus. Indones J Biotechnol 26: 107–114. doi: 10.22146/ijbiotech.65256
Ladics GS, Bannon GA, Silvanovich A, Cressman RF (2007) Comparison of conventional FASTA identity searches with the 80 amino acid sliding window FASTA search for the elucidation of potential identities to known allergens. Mol Nutr Food Res 51: 985–998. doi: 10.1002/mnfr.200600231
Ladics GS, Cressman RF, Herouet-Guicheney C, Herman RA, Privalle L, Song P, Ward JM, McClain S (2011) Bioinformatics and the allergy assessment of agricultural biotechnology products: Industry practices and recommendations. Regul Toxicol Pharmacol 60: 46–53. doi: 10.1016/j.yrtph.2011.02.004
Ladics GS (2018) Assessment of the potential allergenicity of genetically-engineered food crops. J Immunotoxicol 16: 43–53. doi: 10.1080/1547691X.2018.1533904
Lindbo JA, Falk BW (2017) The impact of “coat protein-mediated virus resistance” in applied plant pathology and basic research. Phytopathol 107: 624–634. doi: 10.1094/PHYTO-12-16-0442-RVW
Liu MS, Ko MH, Li HC, Tsai SJ, Lai YM, Chang YM, Wu MT, Chen LFO (2014) Compositional and proteomic analyses of genetically modified broccoli (Brassica oleracea var. italica) harboring an agrobacterial gene. Int J Mol Sci 15: 15188–15209. doi: 10.3390/ijms150915188
Mishra A, Gaur SN, Singh BP, Arora N (2012) In silico assessment of the potential allergenicity of transgenes used for the development of GM food crops. Food Chem Toxicol 50: 1334–1339. doi: 10.1016/j.fct.2012.02.005
Neliana IR, Sawitri WD, Ermawati N, Handoyo T, Sugiharto B (2019) Development of allergenicity and toxicity assessment methods for evaluating transgenic sugarcane overexpressing sucrose–phosphate synthase. Agronomy 9: 23. doi: 10.3390/agronomy9010023
Pali-Schöll I, Untersmayr E, Klems M, Jensen-Jarolim E (2018) The effect of digestion and digestibility on allergenicity of food. Nutrients 10: 1129. doi: 10.3390/nu10091129
Pandey A, Kamle M, Yadava LP, Muthukumar M, Kumar P, Gupta V, Ashfaque M, Pandey BK (2010) Genitically modified food: Its uses, future prospects and safety assesments. Biotechnology 9: 444-458. doi: 10.3923/biotech.2010.444.458
Privalle L, Bannon G, Herman R, Ladics G, McCLain S, Stagg N, Ward J, Herouet-Guicheney C (2011) Heat stability, its measurement, and its lack of utility in the assessment of the potential allergenicity of novel proteins. Regul Toxicol Pharmacol 61: 292–295. doi: 10.1016/j.yrtph.2011.08.009
Rathinam M, Singh S, Pattanayak D, Sreevathsa R (2017) Comprehensive in silico allergenicity assessment of novel protein engineered chimeric Cry proteins for safe deployment in crops. BMC Biotechnology 17: 64. doi: 10.1186/s12896-017-0384-z
Verhoeckx KCM, Vissers YM, Baumert JL, Faludi R, Feys M, Flanagan S, Herouet-Guicheney C, Holzhauser T, Shimojo R, van der Bolt N, Wichers H, Kimber I (2015) Food processing and allergenicity. Food Chem Toxicol 80: 223–240. doi: 10.1016/j.fct.2015.03.005
Widyaningrum S, Pujiasih DR, Sholeha W, Harmoko R, Sugiharto B (2021) Induction of resistance to sugarcane mosaic virus by RNA interference targeting coat protein gene silencing in transgenic sugarcane. Mol Biol Rep 48: 3047–3054. doi: 10.1007/s11033-021-06325-w
Xu JS, Deng YQ, Cheng GY, Zhai YS, Peng L, Dong M, Xu Q, Yang YQ (2019) Sugarcane mosaic virus infection of model plants Brachypodium distachyon and Nicotiana benthamiana. J Integr Agric 18: 2294–2301. doi: 10.1016/S2095-3119(19)62572-4
Yao W, Ruan M, Qin L, Yang C, Chen R, Chen B, Zhang M (2017) Field performance of transgenic sugarcane lines resistant to sugarcane mosaic virus. Front Plant Sci 8: 104. doi: 10.3389/fpls.2017.00104