Document Detail


Heterologous expression of Candida albicans Pma1p in Saccharomyces cerevisiae.
MedLine Citation:
PMID:  23374681     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
Candida albicans is a major cause of opportunistic and life-threatening systemic fungal infections, especially in the immunocompromised. The plasma membrane proton pumping ATPase (Pma1p) is an essential enzyme that generates the electrochemical gradient required for cell growth. We expressed C. albicans Pma1p (CaPma1p) in Saccharomyces cerevisiae to facilitate screening for inhibitors. Replacement of S. cerevisiae PMA1 with C. albicans PMA1 gave clones expressing CaPma1p that grew slowly at low pH. CaPma1p was expressed at significantly lower levels and had lower specific activity than the native Pma1p. It also conferred pH sensitivity, hygromycin B resistance and low levels of glucose-dependent proton pumping. Recombination between CaPMA1 and the homologous non-essential ScPMA2 resulted in chimeric suppressor mutants that expressed functional CaPma1p with improved H(+) -ATPase activity and growth rates at low pH. Molecular models of suppressor mutants identified specific amino acids (between 531-595 in CaPma1p) that may affect regulation of the activity of Pma1p oligomers in S. cerevisiae. A modified CaPma1p chimeric construct containing only 5 amino acids from ScPma2p enabled the expression of a fully functional enzyme for drug screens and structural resolution. © 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.
Authors:
Mikhail V Keniya; Richard D Cannon; Anbình Nguyen; Joel D A Tyndall; Brian C Monk
Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2013-2-4
Journal Detail:
Title:  FEMS yeast research     Volume:  -     ISSN:  1567-1364     ISO Abbreviation:  FEMS Yeast Res.     Publication Date:  2013 Feb 
Date Detail:
Created Date:  2013-2-4     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101085384     Medline TA:  FEMS Yeast Res     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Copyright Information:
© 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.
Affiliation:
The Sir John Walsh Research Institute, School of Dentistry, University of Otago, Dunedin, New Zealand.
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