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Caravel Undergraduate Research Journal

Abstract

Due to its unique ability to serve as both an electron donor and acceptor, iron is utilized as a co-factor for many biological processes, including electron transfer, oxygen binding, and vitamin synthesis. Iron is also a key factor during fungal infections as the human host and invading pathogens battle over limited iron pools. The primary iron-responsive transcription factor Aft1 in the opportunistic pathogenic yeast Candida glabrata responds to iron deficiency by activating expression of iron acquisition genes. However, the mechanisms for sensing intracellular iron levels and regulating Aft1 activity in response to iron are unknown. The C. glabrata iron regulation system shares close homology to a similar system in the non-pathogenic yeast Saccharomyces cerevisiae, in which the monothiol glutaredoxins Grx3/4 and the BolA-like protein Bol2 form [2Fe-2S] binding complexes that deactivate Aft1 under iron replete conditions. To determine whether a similar mechanism controls C. glabrata Af1 activity, we sought to analyze the in vitro interactions between Grx4 and Bol2 from this yeast pathogen. For this project, we successfully subcloned the BOL2 gene from C. glabrata into Escherichia coli overexpression vectors allowing for expression of recombinant Bol2 alone or in complex with its presumed binding partner Grx4. The overexpression conditions identified here will be used in future experiments to purify and characterize the structure and iron regulation function of Bol2 alone or in a complex with Grx4.

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