In Vitro Evaluation of Immunogenicity of Recombinant OMP25 Protein Obtained from Endemic Brucella abortus Biovar 3 as Vaccine Candidate Molecule Against Animal Brucellosis

• Published in: Protein and peptide letters Vol. 28; no. 10; p. 1138
• Main Authors: Atabey, Tuğba, Acar, Tayfun, Derman, Serap, Ordu, Emel, Erdemir, Ayşegül, Taşlı, Pakize Neslihan, Gür, Günseli Kurt, Şahin, Fikrettin, Güllüce, Medine, Arasoğlu, Tülin
• Format: Journal Article
• Language: English
• Published: Netherlands 01.01.2021
• Subjects: Animals; Brucella abortus - genetics; Brucellosis - drug therapy; Cell Survival - drug effects; Cytokines - metabolism; Dose-Response Relationship, Drug; Escherichia coli - chemistry; Escherichia coli - genetics; Humans; Immunogenicity, Vaccine; Membrane Proteins - chemistry; Membrane Proteins - genetics; Membrane Proteins - pharmacology; Mice; Nitric Oxide - metabolism; RAW 264.7 Cells; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - pharmacology; Vaccine Development; Vaccines, Synthetic - chemistry; Vaccines, Synthetic - genetics; Vaccines, Synthetic - pharmacology; OMP25; Brucellosis; fusion protein; vaccine; Brucella abortus; recombinant protein
• ISSN: 1875-5305
• DOI: 10.2174/0929866528666210615104334

Brucellosis is a zoonotic disease that causes serious economic losses due to factors, such as miscarriages and decreased milk yield in animals. Existing live vaccines have some disadvantages, so effective vaccines need to be developed with new technological approaches. The primary objectives of this study were the expression and purification of recombinant Omp25 fusion protein from B. abortus, and the evaluation of the effect of the Omp25 protein on cell viability and inflammatory response. The omp25 gene region was amplified by a polymerase chain reaction and cloned into a Pet102/D-TOPO expression vector. The protein expression was carried out using the prokaryotic expression system. The recombinant Omp25 protein was purified with affinity chromatography followed by GPC (Gel Permeation Chromatography). The MTS assay and cytokine-release measurements were carried out to evaluate cell viability and inflammatory response, respectively. It was determined that doses of the recombinant Omp25 protein greater than 0.1 μg/mL are toxic to RAW cells. Doses of 1 μg/mL and lower significantly increased inflammation due to Nitric Oxide (NO) levels. ELISA results showed that IFN-γ was produced in stimulated RAW 264.7 cells at a dose that did not affect the viability (0.05 μg/mL). However, IL-12, which is known to have a dual role in the activation of macrophages, did not show a statistically significant difference at the same dose. Studies on cell viability and Th1-related cytokine release suggest Omp25 protein to be a promising candidate molecule for vaccine development.