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 Google Scholar
Google Scholar
Right arrow Articles by Upadhya, R.
Right arrow Articles by Weiss, L. M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Upadhya, R.
Right arrow Articles by Weiss, L. M.

 Previous Article  |  Next Article 

Antimicrobial Agents and Chemotherapy, October 2006, p. 3389-3395, Vol. 50, No. 10
0066-4804/06/$08.00+0     doi:10.1128/AAC.00726-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

System for Expression of Microsporidian Methionine Amino Peptidase Type 2 (MetAP2) in the Yeast Saccharomyces cerevisiae{triangledown}

Rajendra Upadhya,1 Hong Shan Zhang,1 and Louis M. Weiss1,2*

Departments of Pathology,1 Medicine, Albert Einstein College of Medicine, Bronx, New York 104612

Received 6 June 2006/ Returned for modification 20 July 2006/ Accepted 8 August 2006

Microsporidia are parasitic protists of all classes of vertebrates and most invertebrates. They recently emerged as important infections in various immunosuppressed and immunocompetent patient populations. They are also important veterinary and agricultural pathogens. Current therapies for microsporidiosis include benzimidazoles, which bind tubulin-inhibiting microtubule assembly, and fumagillin and its derivatives, which bind and inhibit methionine amino peptidase type 2 (MetAP2). Benzimidazoles are not active against Enterocytozoon bieneusi, the most common cause of human microsporidiosis. Fumagillin is active against most microsporidia, including E. bieneusi, but thrombocytopenia has been a problem in clinical trials. There is a pressing need for more-specific microsporidian MetAP2 inhibitors. To expedite and facilitate the discovery of safe and effective MetAP2 inhibitors, we have engineered Saccharomyces cerevisiae to be dependent on Encephalitozoon cuniculi MetAP2 (EcMetAP2) for its growth, where EcMetAP2 is harbored on an episomal uracil-selectable tetracycline-regulated plasmid. We have also constructed a leucine-selectable tetracycline-regulated expression plasmid into which any MetAP2 gene can be cloned. By utilizing a 5-fluoroorotic acid-mediated plasmid shuffle in the EcMetAP2 yeast strain, a yeast strain can be generated whose growth is dependent on MetAP2 from any organism. The level of heterologous MetAP2 gene expression can be controlled by the addition of tetracycline to the growth medium. These yeast strains should permit high-throughput screening for the identification of new inhibitors with high specificity and activity toward microsporidian MetAP2.

* Corresponding author. Mailing address: Albert Einstein College of Medicine, 1300 Morris Park Avenue, Forchhemier Building, Room 504, Bronx, NY 10461. Phone: (718) 430-2142. Fax: (718) 430-8543. E-mail: lmweiss{at}aecom.yu.edu.

{triangledown} Published ahead of print on 17 August 2006.

Antimicrobial Agents and Chemotherapy, October 2006, p. 3389-3395, Vol. 50, No. 10
0066-4804/06/$08.00+0     doi:10.1128/AAC.00726-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.





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