ABSTRACT
An acid phosphatase activity was detected in the supernatant of Haemophilus parasuis, a Gram-negative pleomorphic bacillus and the causative agent of Glässers disease in pigs. To identify the gene responsible for the secreted activity, a genomic library of H. parasuis strain ER-6P was produced in Escherichia coli. Screening of the library allowed identification of two homologs to known phosphatases: PgpB and AphA. PgpB was predicted to be located in the bacterial membrane through six transmembrane domains while AphA was predicted to have a signal peptide. The aphA gene was cloned and expressed in E. coli. Characterization of H. parasuis AphA indicated that this protein belongs to the class B nonspecific acid phosphatases. AphA contained sequence signatures characteristic of this family of phosphatases and its activity was inhibited by EDTA. The optimal pH of recombinant AphA differed from that of the phosphatase activity found in H. parasuis supernatants. In addition, the phosphatase activity from H. parasuis supernatants was not inhibited by EDTA, indicating that H. parasuis AphA does not account for the phosphatase activity observed in the supernatants. Our results demonstrate the presence of a class B acid phosphatase (AphA) in H. parasuis and suggest that the bacterium would also secrete another, as yet unidentified phosphatase (AU)
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Subject(s)
Animals , Acid Phosphatase/isolation & purification , Haemophilus parasuis/immunology , Bacterial Physiological Phenomena/immunology , Swine Diseases/immunology , Biomarkers/analysisABSTRACT
An acid phosphatase activity was detected in the supernatant of Haemophilus parasuis, a Gram-negative pleomorphic bacillus and the causative agent of Glässer's disease in pigs. To identify the gene responsible for the secreted activity, a genomic library of H. parasuis strain ER-6P was produced in Escherichia coli. Screening of the library allowed identification of two homologs to known phosphatases: PgpB and AphA. PgpB was predicted to be located in the bacterial membrane through six transmembrane domains while AphA was predicted to have a signal peptide. The aphA gene was cloned and expressed in E. coli. Characterization of H. parasuis AphA indicated that this protein belongs to the class B nonspecific acid phosphatases. AphA contained sequence signatures characteristic of this family of phosphatases and its activity was inhibited by EDTA. The optimal pH of recombinant AphA differed from that of the phosphatase activity found in H. parasuis supernatants. In addition, the phosphatase activity from H. parasuis supernatants was not inhibited by EDTA, indicating that H. parasuis AphA does not account for the phosphatase activity observed in the supernatants. Our results demonstrate the presence of a class B acid phosphatase (AphA) in H. parasuis and suggest that the bacterium would also secrete another, as yet unidentified phosphatase.
