AurF is a non-heme, iron(II)-dependent monooxygenase. It is known to catalyze the oxidation of arylamines to their respective nitro functional groups. This catalysis is the terminal step in the biosynthesis of a potent antibiotic, aureothin. AurF is also known to have diiron active site that is the epicenter for O2activation. There have been experimental and computational studies examining the mechanism of this metalloenzyme, and the differing metal centers in AurF have been spectroscopically characterized. However, there are still many questions regarding how the change of the metal ions affect the protein’s overall stability. When substituted with manganese(II), this enzyme has also been shown to maintain its reactivity. Therefore, to study AurF’s thermostability and provide further characterization, an array of differential scanning calorimetry (DSC) experiments will be performed. In a DSC experiment, we will thermally denature the enzyme and measure the changes in melting temperature (Tm) and heat capacity (ΔCp) of both the manganese and iron forms of AurF. Using this data, conformational changes can be determined and compared for the two metals ions forms of AurF, giving insight into their effect on globular protein stability. AurF’s bimetallic center makes it an interesting molecule for study, and this research hopes to give a more detailed look into the stability and characterization of AurF, and shed more light on its potential use in biosynthesis.

Using differential scanning calorimetry to study the thermostability of the diiron metalloenzyme AurF

2019 Fall National Meeting

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