Download Search for the potential antigens present in the outer membrane of

Search for the potential antigens present in the outer membrane of
Pasteurella multocida by immunoproteomics
Hsiao-Ping Chou1, Keng-Hung Liu1, Cheng-Hao Huang1, Wei-Chen Yang1,
Ya-Sheng Yu1, Chia-Ching Lin1, Yeou-Guang Tsay3* Ting-Feng Wu1,2*
1
3
Department of biotechnology and 2Research Center of Biotechnology, Southern
Taiwan University, Tainan, Taiwan
Institute of Biochemistry and Molecular Biology, National Yang Ming University,
Taipei, Taiwan
Gram-negative coccobacillus pathogen which is Pasteurella multocida result in
fowl cholera and hemorrhagic septicaemia. Current vaccines against fowl cholera
including the live naturally attenuated and the killed whole cells, provide only limited
protection. In this study, we tried to find the possible antigens in outer membrane,
which may serve as the potential subunit vaccines. Two dimensional (2-D) western
blotting with the elicited-sera, which were taken from the formalin-killed
bacteria-administrated mice was exploited to found out the possible antigens present
in the outer membrane of P. multocida. Western blotting demonstrated that 23 proteins
were able to interact with the sera. These 23 proteins were further identified using
high performance liquid chromatography electro-spray tandem mass spectrometry.
One of these 23 proteins was recognized as pm1992 which is an outer membrane
protein and other recognized proteins were involved in energy metabolism. Owing to
pm1992 protein’s specific membrane location and unknown function, we anticipated
that pm1992 may be a possible subunit vaccine candidate. Therefore, pm1992 gene
was cloned into the pPAL7 procaryotic expression vector and purified with Profinity
eXact™ Protein Purification System (Bio-Rad). The results of western blotting
showed that the purified pm1992 could be recognized by induced-sera, indicating that
pm1992 protein may be a potential subunit vaccine candidate.