Haa1
This project aims the structural characterization of the Yeast transcription factor (TF) Haa1p and its complex with the Haa1p-responsive element (HRE). Haa1p was demonstrated to control a novel acetic acid-sensing system in yeast. This signalling pathway was also found critical for yeast resistance to lactic acid. Acetic and lactic acids are potent fungistatic agents used frequently by food and beverages industries as preservatives. The understanding of the mechanisms underlying the yeast response that counteract the deleterious effects of organic acids is essential to guide suitable preservation strategies. Moreover, acetic and lactic acids are also bypass products of alcoholic fermentation. Their accumulation in the growth medium reaching toxic levels, often leads to decreased yields and stuck fermentations. Results emerging from this project may lead to the engineering of more robust industrial yeast strains. The here proposed structural characterization will involve a genetic approach, through the production of site-directed amino-acid mutants, a crystallographic approach, through the crystal structural determination of Haa1p, its DNA binding domain and their complexes with HRE, and the analysis of Haa1p conformational alterations by weak acid induction using fluorescence techniques in vitro, with Thermal Shift Assays (TSA), and in vivo, with Fluorescence Resonance Energy Transfer (FRET). The combination of these strategies will enable the elucidation of the crucial residues for DNA binding activity and allow the comparison of the tridimensional structures of functional versus non-functional Haa1-DNA complexes.
Project webpage Start year 01/05/2016 End year 31/10/2019 ID PTDC/BBB-BEP/0385/2014 iBB Role Partner iBB Budget 120 926,00 € Research Group BSRG PI Isabel Sá Correia Project Partners ITQB Status Completed Funding FCT