This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Schmidt, J. A.
Right arrow Articles by Markham, P. F.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Schmidt, J. A.
Right arrow Articles by Markham, P. F.

 Previous Article  |  Next Article 

Journal of Bacteriology, September 2004, p. 5790-5798, Vol. 186, No. 17
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.17.5790-5798.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Mycoplasma hyopneumoniae p65 Surface Lipoprotein Is a Lipolytic Enzyme with a Preference for Shorter-Chain Fatty Acids

Jono A. Schmidt, Glenn F. Browning,* and Philip F. Markham

Department of Veterinary Science, Veterinary Preclinical Centre, The University of Melbourne, Parkville, Victoria, Australia

Received 10 March 2004/ Accepted 19 May 2004

Mycoplasma hyopneumoniae is the most significant bacterial pathogen of the respiratory tract of swine. p65 is an immunodominant surface lipoprotein of M. hyopneumoniae that is specifically recognized during disease. Analysis of the translated amino acid sequence of the gene encoding p65 revealed similarity to the GDSL family of lipolytic enzymes. To examine the lipolytic activity of p65, the gene was cloned and expressed in Escherichia coli after truncation of the prokaryotic lipoprotein signal sequence and mutagenesis of the mycoplasma TGA tryptophan codons. After treatment with thrombin, the recombinant glutathione S-transferase (GST)-p65 protein yielded a 66-kDa fusion protein cleavage product corresponding in size to the mature p65 protein. The esterase activity of recombinant GST-p65 was indicated by the formation of a cleared zone on tributyrin agar plates and the hydrolysis of p-nitrophenyl esters of caproate (pNPC) and p-nitrophenyl esters of palmitate (pNPP). Lipase activity was indicated by the hydrolysis of the artificial triglyceride 1,2-O-dilauryl-rac-glycero-3-glutaric acid resorufin ester. Using pNPC and pNPP as substrates, recombinant GST-p65 had optimal activity between pHs 9.2 and 10.2 and at a temperature higher than 39°C. Calcium ions did not increase the activity of recombinant GST-p65. Rabbit anti-p65 antibodies inhibited the activity of recombinant GST-p65 and also inhibited the growth of M. hyopneumoniae in vitro. Examination of the kinetic parameters of recombinant GST-p65 for the hydrolysis of pNPC and pNPP indicated a preference for the shorter fatty acid chain of pNPC. The physiological and/or pathogenic role of mycoplasma lipolytic enzymes has not been determined, but they are likely to play an important role in mycoplasmas' nutritional requirements for long-chain fatty acids and may reduce the function of lung surfactants in mycoplasma-induced respiratory diseases. This is the first report of the lipolytic activity of a lipid-modified surface immunogen of a mycoplasma.

* Corresponding author. Mailing address: Department of Veterinary Science, Veterinary Preclinical Centre, The University of Melbourne, Parkville, Victoria 3010, Australia. Phone: 61 3 8344 7342. Fax: 61 3 8344 7374. E-mail: glenfb{at}unimelb.edu.au.

Journal of Bacteriology, September 2004, p. 5790-5798, Vol. 186, No. 17
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.17.5790-5798.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Gil, F., Catalao, M. J., Moniz-Pereira, J., Leandro, P., McNeil, M., Pimentel, M. (2008). The lytic cassette of mycobacteriophage Ms6 encodes an enzyme with lipolytic activity. Microbiology 154: 1364-1371 [Abstract] [Full Text]  
  • Schmidt, J. A., Browning, G. F., Markham, P. F. (2007). Mycoplasma hyopneumoniae mhp379 Is a Ca2+-Dependent, Sugar-Nonspecific Exonuclease Exposed on the Cell Surface. J. Bacteriol. 189: 3414-3424 [Abstract] [Full Text]  
  • Ruiz, C., Falcocchio, S., Pastor, F. I. J., Saso, L., Diaz, P. (2007). Helicobacter pylori EstV: Identification, Cloning, and Characterization of the First Lipase Isolated from an Epsilon-Proteobacterium. Appl. Environ. Microbiol. 73: 2423-2431 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»