Alerjen Proteinlerin Saflaştırılmasında Kullanılan Moleküler Yöntemler

Yıl 2018, Cilt: 4 Sayı: 2, 13 - 28, 12.09.2018

Serkan Sugeçti Adem İmalı Ferudun Koçer

Öz

Özet

İnsanların % 10 unda rastlanan alerjenite
yurtdışında birçok araştırıcı tarafından farklı yöntemler ile belirlenmektedir.
Bu çalışmada, alerjen proteinlerinin saflaştırılması için kullanılan yöntemler
incelenmiştir.

Alerjenlerin belirlenmesinde
kullanılan tamponlar ve işlemler birçok farklılıklar içermektedir. İncelenen
kromatografik yöntemler; jel filtrasyon kromatografisi, iyon değişim kromatografisi,
affinite kromatografisi, ters faz kromatografisi, hidrofobik etkileşim kromatografisi,
membran kromatografisi, yüksek performans sıvı kromatografisi (HPLC) ve
elektroforetik yöntemler ise; izoelektrik odaklama, sodyum dodesil sülfat
poliakrilamid jel elektroforezi (SDS-PAGE) ve 2 boyutlu elektroforezdir.

Kullanılacak olan yöntemler
saflaştırılması arzu edilen proteinin fiziksel, kimyasal ve biyolojik
özellikleri göz önüne alınarak seçilmektedir. Proteini, en saf halde elde
edebilmek için bu yöntemlerin çoğu ardışık olarak uygulanmaktadır. En fazla
miktarda protein, membran kromatografisiyle, en az miktarda protein, affinite
kromatografisi ile elde edilmektedir. Bu sebeple birçok yöntemin kendine özgü
özellikleri ve avantajları mevcuttur.

Güncel bilgiler ile kullanılan
kimyasal ve cihazların belirlenmesi ve uygun yöntemler ile ortak bir kullanım
tespiti birçok avantajlar sağlayacaktır. Son yıllarda, elde edilen saf
proteinler, birçok alanda kullanıma sunulmakta ve bu alan hızla gelişen bir
sektör olarak diğer sektörler arasında yerini almaktadır.  Saflaştırmada
izlenecek kriterler incelenerek alerjenlerin saflaştırılmasında yarar
sağlayacağı düşünülmektedir.

Anahtar Kelimeler: Alerjen, saflaştırma, elektroforetik yöntemler, kromatografik yöntemler

 

 

 

Molecular Methods Used in the Purification of Allergen Proteins

Abstract

Allergenicity of 10% of people is
determined by different methods by many researchers abroad. In this study, the
methods used to purify the protein allergen is examined.

Buffers and procedures used to
identify allergens include many variationsExamination of chromatographic
methods; gel filtration chromatography, ion exchange chromatography, affinity
chromatography, reversed phase chromatography, hydrophobic interaction
chromatography, membrane chromatography, high performance liquid chromatography
(HPLC) and electrophoretic methods; isoelectric focusing, sodium dodecyl
sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and 2-D electrophoresis.

The methods to be used are selected
by considering the physical, chemical and biological properties of the desired
protein. Most of these methods are applied sequentially to obtain the protein
in its purest form. The maximum amount of protein is obtained by membrane
chromatography, with minimal amount of protein, affinity chromatography. For this
reason, many methods have their own characteristics and advantages.

Identification of the current
information, the chemicals and devices used, and the determination of a common
usage by appropriate methods will provide many advantages. In recent years, the
obtained pure proteins have been used in many fields and this area is among the
other sectors as a rapidly developing sector. It is thought that the purifying
criteria will be useful for the purification of allergens.

Keywords: Alergen,
purification, electrophoretic methods, chromatographic methods

Anahtar Kelimeler

Alerjen, saflaştırma, elektroforetik yöntemler, , kromatografik yöntemler

Kaynakça

• (1) Aas K, Bachert C, Bergmann K, Bergmann R, Bonını S, Bousqet J, De Weck A, Farkas I, Hejdenberg K.. European White Paper: Allergic Diseases as a Health Problem. The UCB Institute of Allergy. Brussels, Belgium. 1997
• (2) Huynen M, Menne BC. Phenology and Human Health: Allergic Disorders, Report of a WHO meeting in Rome, Italy. 16–17 Jan 2003. Health and Global Environmental Series EUR/03/5036791. World Health Organization, 2003; 64.
• (3) Puc M. Characterization of Pollen Allergens. Annals of Agricultural and Environmental Medicine, 2003; 10: 143-149.
• (4) Sin A B, Pınar NM, Mısırlıgil Z, Çeter T, Yıldız A, Alan Ş. Polen Allerjisi; Türkiye Allerjik Bitkilerine Genel Bir Bakış. 1.Baskı, Engin Yayınevi, Ankara. 2007.
• (5)Pastorello EA, Trambaioli C. Isolation of food allergens. J Chromatogr B Biomed Sci Appl 2001; 756(1–2):71–84
• (6) Scheiner O, Kraft D. Basic and Practical Aspects of Recombinant Allergens. Allergy, 1995; 50(5):384–391.
• (7) Thomas WR. Mite Allergens Groups I-VII. A Catalogue of Enzymes. Clinical & Experimental Allergy, 1993; 23(5): 350–353.
• (8) Chapman MD.. Allergen Nomenclature. In: Lockey RF, Bukantz SC, Bousquet J, editors. Allergens and Allergen Immunotherapy. New York: Marcel Decker, 2003;51-64.
• (9) Chapman MD.. Indoor Allergens, Diagnosis and Treatment of Allergic Disease, Chapter 25, 2010; 266-273.(10) Cacciola RR, Sarva M, Polosa R.. Adverse Respiratory Effects and Allergic Susceptibility in Relation to Particulate Air Pollution: Flirting With Disaster. Allergy, 2002; 57: 281-6.
• (11) D’amato G. Environmental Urban Factors (Air Pollution and Allergens) and The Rising Trends in Allergic Respiratory Diseases. Allergy, 2002; 57: 30-3.
• (12) Pınar NM, Akgül G, Tuğ G N. Palinoloji Laboratuar Kılavuzu, Ankara Üniversitesi Yayınları, No: 66 Ankara, 2003; 71.
• (13) Chapman MD, Pomes A, Breiteneder H, Ferreira F. Nomenclature and Structural Biology of Allergens. Journal of Allergy and Clinical Immunology, 2007; 119: 414-420.
• (14) Çeter T, Pinar NM. Türkiye'de Yapılan Atmosferik Fungus Spor Çalışmaları ve Kullanılan Yöntemler. Asthma Allergy Immunology/Astim Allerji Immunoloji, 2009; 7(1).
• (15) İmalı A, Koçer F, Yalçınkaya B. Microfungus Flora of Indoor and Outdoor Air in Primary Schools, Çorum, Turkey. Australian Journal of Basic and Applied Sciences (AJBAS), 2011; 2274-2278.
• (16) Unger A, Stöger P, Simon-Nobbe B, Susani M, Crameri R, Ebner C, Hintner H, Breitenbach M. Clinical Testing of Recombinant Allergens of The Mold Alternaria alternata. International Archives of Allergy and Immunology, 1999; 118: 220-221.
• (17) Bush RK, Prochnau JJ. Alternaria-induced Asthma. Journal of Allergy and Clinical Immunology, 2004; 113(2), 227-234.
• (18) Gasilova N, Girault HH. Component-Resolved Diagnostic of Cow’s Milk Allergy by Immunoaffinity Capillary Electrophoresis–Matrix Assisted Laser Desorption/Ionization Mass Spectrometry. Analytical Chemistry, 2014; 6:6337-6345.
• (19) King TP, Hoffman D, Lowenstein H, Marsh DG, Platts-Mills TA, Thomas W. Allergen Nomenclature. Bulletin of the World Health Organization, 1994; 72:797–806
• (20) Esch ER. Allergen Source Materials and Quality Control of Allergenic Extracts, A Companion to Methods in Enzymology 1997; 13, 2–13.
• (21) Aalberse RC. Structural Biology of Allergens. Journal of Allergy and Clinical Immunology, 2000; 106(2):228–238.
• (22) Burgess RR, Deutscher MP. (Eds.). Guide to Protein Purification (Vol. 463). Academic Press. 2009; 854
• (23) Towbin H, Staehelin T, Gordon J. Electrophoretic Transfer of Proteins From Polyacrylamide Gels to Nitrocellulose Sheets: Procedure and Some Applications. Proceedings of the National Academy of Sciences, 1979; 76: 4350-4.
• (24) Raftery MJ, Saldanha RG, Geczy CL, Kumar RK. Mass Spectrometric Analysis of Electrophoretically Separated Allergens and Proteases in Grass Pollen Diffusates. Respiratory Research, 2003; 4(1), 10.
• (25) Kao SH, Su SN, Huang SW, Tsai JJ, Chow LP. Sub‐Proteome Analysis of novel IgE‐binding proteins from Bermuda Grass Pollen. Proteomics, 2005; 5(14):3805-3813.
• (26) Petersen A, Suck R, Lindner B, Georgieva D, Ernst M, Notbohm H, Wicklein D, Cromwell, O, Becker WM. Phl p 3: Structural and Immunological Characterization of A Major Allergen of Timothy Grass Pollen. Clinical & Experimental Allergy, 2006;36(6): 840-849.
• (27) Erler A, Hawranek T, Krückemeier L, Asam C, Egger M, Ferreira F, Briza P. Proteomic Profiling of Birch (Betula verrucosa) Pollen Extracts From Different Origins. Proteomics, 2011; 11(8):1486-1498.
• (28) Koçer F. Betulaceae Familyası Duyarlı Hastalarda Alnus orientalis (Doğu Kızılağacı) Decne. Polenine Karşı Gösterilen IgE Reaktivite Profilinin Belirlenmesi ve Majör Alerjenlerinin Saflaştırılması, Bülent Ecevit Üniversitesi, Fen Bilimleri Enstitüsü, 2015; s.93.
• (29) Ghosh N, Sircar G, Saha B, Pandey N, Bhattacharya SG. Search for Allergens From The Pollen Proteome of Sunflower (Helianthus annuus L.): a Major Sensitizer for Respiratory Allergy Patients. PloS one, 2015; 10(9), e0138992.
• (30) Bouley J, Groeme R, Le Mignon M, Jain K, Chabre H, Bordas-Le Floch V, Baron-Bodo V, Lombardi V, Mascarell L, Batard T, Nony E, Moingeon P. Identification of The Cysteine Protease Amb a 11 as a Novel Major Allergen From Short Ragweed. Journal of Allergy and Clinical Immunology, 2015; 136(4), 1055-1064.
• (31) Kılıç E, Köseoğlu F, Yılmaz H. Enstrümental Analiz İlkeleri. Bilim Yayıncılık, ISBN: 975-55-041-6, 1997; 849
• (32) Tuncer M. Protein Saflaştırma 1: Kromatografik Teknikler, Mersin Üniversitesi Yayınları, ISBN: 978-9756-900215, 2008; 268.
• (33) Snyder LR, Kirkland JJ, Dolan JW. Introduction to Modern Liquid Chromatography. John Wiley & Sons. 2010; 912.
• (34) Riley CM, Rosanske TW, Riley SRR. Specification of Drug Substances and Products: Development and Validation of Analytical Methods 2013; Vol. 3. Newnes
• (35) Toker NY. Protein Saflaştırılması İle İlgili Bazı Metodlar, Vetfakdergi, 2000;2:12.
• (36) Vijayalakshmi MA. Biochromatography Theory and Practice, Taylor and Francis London & NY. 2002
• (37) Metin K. Moleküler Biyoloji Teknikleri II: Protein Analiz Teknikleri. Moleküler Biyoloji (Yıldırım, A., Bardakcı, F., Karataş, M. ve Tanyolaç, B.), Nobel Yayıncılık, Ankara, 2007; 555-598,
• (38) Mourey TH. SEC Molecular-Weight-Sensitive Detection. International Journal of Polymer Analysis and Characterization, 2004; 9: 97-135.
• (39) Suzuki M, Komiyama N, Itoh M, Itoh H, Sone T, Kino K, Takagi I, Ohta N. Purification, Characterization and Molecular Cloning of Cha o 1, a Major Allergen of Chamaecyparis obtusa (Japanese cypress) Pollen. Molecular Immunology, 1996; 33(4):451-460.
• (40) Tejera ML, Villalba M, Batanero E, Rodríguez R. Identification, Isolation, and Characterization of Ole e 7, a New Allergen of Olive Tree Pollen. Journal of Allergy and Clinical Immunology, 1999; 104(4), 797-802.
• (41)Cadot P, Diaz JF, Proost PJVD, Van Damme J, Engelborghs Y, Stevens EAM, Ceuppens JL. Purification and Characterization of an 18-kDa Allergen of Birch (Betula verrucosa) Pollen: Identification as a Cyclophilin. Journal of Allergy and Clinical Immunology, 2000; 105(2), 286-291.
• (42) Huecas S, Villalba M, Rodrı́guez R.. Ole e 9, a Major Olive Pollen Allergen is a 1, 3-β-glucanase Isolation, Characterization, Amino Acid Sequence, and Tissue Specificity. Journal of Biological Chemistry, 2001; 276(30):27959-27966.
• (43) Wu WC, Tam MF, Peng HJ, Tsai LC, Chi CW, Chang ZN. Isolation and Partial Characterization of a 46-kd Allergen of Bermuda Grass Pollen. Journal of Biomedical Science, 2001; 8(4): 342-348.
• (44)Calabozo B, Barber D, Polo F. Purification and Characterization of The Main Allergen of Plantago lanceolata Pollen, Pla l 1. Clinical & Experimental Allergy, 2001; 31(2): 322-330.
• (45) Ibarrola I, Arilla MC, Martínez A, Asturias JA. Identification of a Polygalacturonase as a Major Allergen (Pla a 2) from Platanus acerifolia pollen. Journal of Allergy and Clinical İmmunology, 2004; 113(6):1185-1191.
• (46) Barderas R, Villalba M, Pascual CY, Batanero E, Rodríguez R. Profilin (Che a 2) and Polcalcin (Che a 3) are Relevant Allergens of Chenopodium album Pollen: Isolation, Amino Acid Sequences, and Immunologic Properties. Journal of Allergy and Clinical Immunology, 2004; 113(6):1192-1198.
• (47) Movérare R, Everberg H, Carlsson R, Holtz A, Thunberg R, Olsson P, Brostedt P, Högbom E. Purification and Characterization of The Major Oak Pollen Allergen Que a 1 for Component-Resolved Diagnostics Using ImmunoCAP®. International Archives of Allergy and Immunology, 2008; 146(3):203-211.
• (48) Planque M, Arnould T, Dieu M, Delahaut P, Renard P, Gillard N. Advances in Ultra-High Performance Liquid Chromatography Coupled to Tandem Mass Spectrometry For Sensitive Detection of Several Food Allergens in Complex and Processed Foodstuffs. Journal of Chromatography A, 2016; 1464:115-123.