Neuraminidase production by a Pasteurella haemolytica A1 strain associated with bovine pneumonia

• Published in: Infection and Immunity 1993-01, Vol.61 (1), p.253-259
• Main Authors: Straus, D.C. (Texas Tech University Health Sciences Center, Lubbock, TX), Unbehagen, P.J, Purdy, C.W
• Format: Article
• Language: English
• Subjects: Animals; Bacteriology; Biological and medical sciences; Cattle; Cattle Diseases - microbiology; Cell Division; Chromatography, Gel; Chromatography, Ion Exchange; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; GLICOSIDASAS; Glycoproteins - metabolism; GLYCOSIDASE; Hot Temperature - adverse effects; Hydrogen-Ion Concentration; Mannheimia haemolytica - enzymology; Mannheimia haemolytica - physiology; Metabolism. Enzymes; Microbiology; Molecular Weight; Neuraminidase - biosynthesis; Neuraminidase - isolation & purification; PASTEURELLA; Pasteurella haemolytica; Pneumonia - microbiology; Pneumonia - veterinary; Substrate Specificity
• ISSN: 0019-9567; 1098-5522
• DOI: 10.1128/IAI.61.1.253-259.1993

The properties of an extracellular neuraminidase produced by a Pasteurella haemolytica A1 strain (isolated from a case of bovine pneumonia) during growth in a defined medium were examined in this investigation. This enzyme, isolated from concentrated culture supernatants of P. haemolytica A1, was active against N-acetylneuramin lactose, human alpha 1-acid glycoprotein, fetuin, and bovine submaxillary mucin. Neuraminidase production paralleled bacterial growth in a defined medium and was maximal in the stationary phase of growth. The enzyme was purified to homogeneity by a combination of salt fractionation, ion-exchange chromatography on DEAE-Sephacel, and gel filtration on Sephadex G-200. These procedures yielded an enzyme preparation that possessed a specific activity of 100.62 micromol of sialic acid released per min per mg of protein against human alpha 1-acid glycoprotein. The Km value for this enzyme with human alpha 1-acid glycoprotein as the substrate was 1.1 mg/ml, and the enzyme possessed a pH optimum of 6.5. The P. haemolytica A1 neuraminidase had a molecular weight of approximately 150,000 as estimated by gel filtration and approximately 170,000 when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme was stable at 4 degrees C for 3 h. At 37 degrees C for 3 h, 25% of enzymatic activity was lost. Approximately 55% of the enzyme activity was lost within 30 min at 50 degrees C, with greater than 70% of the enzyme activity being destroyed within 10 min at temperatures of greater than 65 degrees C