Document Detail

A high-throughput assay for modulators of NNT activity in permeabilized yeast cells.
MedLine Citation:
PMID:  21602486     Owner:  NLM     Status:  MEDLINE    
Nicotinamide nucleotide transhydrogenase (NNT) mutant mice show glucose intolerance with impaired insulin secretion during glucose tolerance tests. Uncoupling of the β cell mitochondrial metabolism due to such mutations makes NNT a novel target for therapeutics in the treatment of pathologies such as type 2 diabetes. The authors propose that increasing NNT activity would help reduce deleterious buildup of reactive oxygen species in the inner mitochondrial matrix. They have expressed human Nnt cDNA for the first time in Saccharomyces cerevisiae, and transhydrogenase activity in mitochondria isolated from these cells is six times greater than is seen in wild-type mitochondria. The same mitochondria have partially uncoupled respiration, and the cells have slower growth rates compared to cells that do not express NNT. The authors have used NNT's role as a redox-driven proton pump to develop a robust fluorimetric assay in permeabilized yeast. Screening in parallel a library of known pharmacologically active compounds (National Institute of Neurological Disorders and Stroke collection) against NNT ± cells, they demonstrate a robust and reproducible assay suitable for expansion into larger and more diverse compound sets. The identification of NNT activators may help in the elucidation of the role of NNT in mammalian cells and assessing its potential as a therapeutic target for insulin secretion disorders.
Nicholas A Meadows; Barbara Saxty; Mary S Albury; Catherine A Kettleborough; Frances M Ashcroft; Anthony L Moore; Roger D Cox
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2011-05-20
Journal Detail:
Title:  Journal of biomolecular screening     Volume:  16     ISSN:  1552-454X     ISO Abbreviation:  J Biomol Screen     Publication Date:  2011 Aug 
Date Detail:
Created Date:  2011-07-26     Completed Date:  2011-12-12     Revised Date:  2014-02-24    
Medline Journal Info:
Nlm Unique ID:  9612112     Medline TA:  J Biomol Screen     Country:  United States    
Other Details:
Languages:  eng     Pagination:  734-43     Citation Subset:  IM    
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MeSH Terms
Enzyme Activation / drug effects
Gene Expression Regulation, Fungal
High-Throughput Screening Assays*
Mitochondria / genetics,  metabolism
NADP Transhydrogenases / genetics,  metabolism*
Reproducibility of Results
Saccharomyces cerevisiae / drug effects*,  genetics,  growth & development,  metabolism*
Small Molecule Libraries / pharmacology
Grant Support
084655//Wellcome Trust; MC_G1000806//Medical Research Council; MC_U142661184//Medical Research Council; //Medical Research Council; //Wellcome Trust
Reg. No./Substance:
0/Small Molecule Libraries; EC 1.6.1.-/NADP Transhydrogenases

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