1. Biotin protein ligase from Candida albicans: Expression, purification and development of a novel assay
- Author
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John C. Wallace, Nicole R. Pendini, Lisa M. Bailey, Steven W. Polyak, Grant W. Booker, Matthew C.J. Wilce, Pendini, Nicole R, Bailey, Lisa M, Booker, Grant W, Wilce, Matthew CJ, Wallace, John C, and Polyak, Steven W
- Subjects
assay development ,Antifungal Agents ,Drug Evaluation, Preclinical ,Biophysics ,Biotin ,Biology ,Biochemistry ,Fungal Proteins ,chemistry.chemical_compound ,Candida albicans ,Carbon-Nitrogen Ligases ,biotin protein ligase ,Enzyme Inhibitors ,Molecular Biology ,Pyruvate Carboxylase ,chemistry.chemical_classification ,Fungal protein ,DNA ligase ,Fatty Acids ,Gluconeogenesis ,Acetyl-CoA carboxylase ,biology.organism_classification ,fungicide, drug discovery ,Molecular biology ,Recombinant Proteins ,Protein Structure, Tertiary ,Pyruvate carboxylase ,Metabolic pathway ,Enzyme ,chemistry ,protein structure and function ,candida albicans ,Biological Assay ,Acetyl-CoA Carboxylase - Abstract
Biotin protein ligase (BPL) is an essential enzyme responsible for the activation of biotin-dependent enzymes through the covalent attachment of biotin. In yeast, disruption of BPL affects important metabolic pathways such as fatty acid biosynthesis and gluconeogenesis. This makes BPL an attractive drug target for new antifungal agents. Here we report the cloning, recombinant expression and purification of BPL from the fungal pathogen Candida albicans. The biotin domains of acetyl CoA carboxylase and pyruvate carboxylase were also cloned and characterised as substrates for BPL. A novel assay was established thereby allowing examination of the enzyme's properties. These findings will facilitate future structural studies as well as screening efforts to identify potential inhibitors. © 2008 Elsevier Inc. All rights reserved. Refereed/Peer-reviewed
- Published
- 2008