Turk J Immunol. 2018; 6(1): 8-15 | DOI: 10.25002/tji.2018.698

Identification of Hemagglutinin Protein from Streptococcus pneumoniae Pili as a Vaccine Candidate by Proteomic Analysis

Diana Chusna Mufida¹, Kusworini Handono², Sumarno Reto Prawiro³, Sanarto Santoso^3
1Faculty of Medicine, University of Jember, Jember, East Java Indonesia, Laboratory of Microbiology, Jember/East Java, Indonesia
²Faculty of Medicine, University Brawijaya, Malang, East Java, Indonesia, Laboratory of Pathology Clinic, Malang/East Java, Indonesia
³Faculty of Medicine, University Brawijaya, Malang, East Java, Indonesia, Laboratory of Microbiology, Malang/East Java, Indonesia

Introduction: Streptococcus pneumoniae is a Gram-positive, bacterial human pathogen that causes a large number of respiratory tract infections, such as pneumonia and sinusitis, as well as invasive diseases such as septicemia and meningitis. Among the surface-exposed virulence factors, the pneumococcal pilus has been shown to play a role in host-cell adhesion. Hemagglutinin activity has been shown to correlate with adhesion ability. The purpose of this study was to identify of S. pneumoniae pili proteins with hemagglutinin activity, and to proteonomically analyze such proteins as potential vaccine candidates. Materials and Methods: The purification of pili was carried out by a pilus cutter method. After SDS-PAGE separation, pili proteins were purified by electroelution and dialysis. After a hemagglutinination assay with mouse erythrocytes, the hemagglutinin protein was identified with Liquid Chromatography/Mass Spectrometry, along with assessments of antigenicity and epitope mapping. Results: SDS-PAGE showed that pili protein had major bands of molecular weights of 67, 54, 25, and 11 kDa. The 54 kDa pili protein was the hemagglutinin. Computational bioinformatics analysis by Mascot Server revealed this hemagglutinin matched with the Rrgb protein of S. pneumoniae (gi: 299856655). Conclusion: The analysis of its antigenicity, and epitope mapping showed that a novel protein was identified as the pilus backbone (Rrgb) of Streptococcus pneumoniae, and has a high antigenicity suitable for further development as a vaccine candidate.

Keywords: Streptococcus pneumoniae, hemagglutinin, pili, proteomic analysis

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Identification of Hemagglutinin Protein from Streptococcus pneumoniae Pili as a Vaccine Candidate by Proteomic Analysis

Diana Chusna Mufida¹, Kusworini Handono², Sumarno Reto Prawiro³, Sanarto Santoso^3
1Faculty of Medicine, University of Jember, Jember, East Java Indonesia, Laboratory of Microbiology, Jember/East Java, Indonesia
²Faculty of Medicine, University Brawijaya, Malang, East Java, Indonesia, Laboratory of Pathology Clinic, Malang/East Java, Indonesia
³Faculty of Medicine, University Brawijaya, Malang, East Java, Indonesia, Laboratory of Microbiology, Malang/East Java, Indonesia

Giriþ: Streptococcus pneumoniae, pnömoni ve sinüzit gibi solunum yolu infeksiyonu olduðu kadar septisemi ve menenjit gibi daha ciddi infeksiyonlara da yol açan Gram-pozitif bir bakteridir. Virulansý belirleyen yüzey ile iliþkili öðeleri içinde, pnömokok pilusu önemli bir rol oynar. Hemaglütünin aktivitesinin ise, bakterinin yapýþma özelliði ile iliþkili olduðu gösterilmiþtir ve bu proteinlerin proteom ile irdelenmesi, oluþturulabilecek bir aþýda kullanýlabilmelerini saðlayabilir. Gereçler ve Yöntemler: Piluslarýn saflaþtýrýlmasý, piluslarýn kesilmesi ile yapýlmýþtýr. Pili proteini SDS-PAGE kullanarak, elektroelusyon ve diyaliz ile saflaþtýrýlmýþtýr. Fare eritrositleri ile yapýlan hemaglütinasyon testinden sonra hemaglütinin proteini, epitop haritalamasý ve antijenik özelliði Sývý Kromatografisi ve Kitle Spektrometresi ile tanýmlandý. Bulgular: SDS-PAGE yöntemi ile pili proteinleri ana bantlarý 67, 52, 25 ve 11kDA moleküler aðýrlýðýnda saptandý. 54 KDa aðýrlýðýndaki pilus proteini hemaglütinin antijeni idi. Mascot sunucusu ile yapýlan hesaplamalý biyobilgi analizi bu hemaglutininin S. pneumoniae’inin Rrgb proteini ile eþleþtiðini gösterdi. Sonuç: Antijen özelliði ve epitop haritalama Streptococcus pneumoniae’dan pilus omurgasý (Rrgb) olarak tanýmlanan bu yeni proteinin yüksek antijenik özelliðinden dolayý aþý geliþtirmede uygun bir aday olabileceðini göstermektedir.

Anahtar Kelimeler: Streptococcus pneumoniae, hemagglutinin, pili, proteomik analiz

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