Semester of Graduation

spring 2022


Master of Science (MS)


Biological Sciences

Document Type



Iron-sulfur proteins are ubiquitous in nature, and they play a crucial role in multifarious biological processes such as electron transfer, enzymatic catalysis, and homeostatic regulation. MitoNEET is the first discovered iron-sulfur protein in the outer mitochondrial membrane. Previous studies have shown that the [2Fe-2S] clusters of mitoNEET can be reduced by FMNH2 in the presence of NADH and Flavin reductase (Fre) under aerobic conditions suggesting that mitoNEET is novel redox enzyme catalyzing electron transfer from NADH via FMN to oxygen.

The molecular docking models suggest that mitoNEET has a specific binding site for FMN close to its [2Fe-2S] cluster in each monomer. Since the closest distance between the bound FMN and the [2Fe-2S] cluster is approximately 10 Å, it may promote rapid electron transfer from FMNH2 to the [2Fe-2S] clusters. In the first part of this work, we have explored the FMN binding site in mitoNEET utilizing two FMN analogs: lumiflavin and lumichrome. We found that Lumiflavin, as FMN, can reduce the mitoNEET [2Fe-2S] clusters in the presence of NADH and Fre under aerobic conditions. Additionally, lumiflavin can change EPR spectrum of the reduced mitoNEET [2Fe-2S] clusters and under blue light exposure form a covalently bound complex with mitoNEET. Lumichrome, instead, inhibits FMNH2 mediated electron transfer activity indicating lumichrome’s potential as an antagonist of electron transfer activity of mitoNEET.

Ubiquinone-10 is a unique electron carrier found in both inner and outer mitochondrial membrane. Ubiquinone-10 accepts electrons from NADH and FADH2 of several different redox pathways. In the second part of this work, we investigated the oxidation kinetics of the reduced [2Fe-2S] clusters of mitoNEET with a fusion protein, YneM-mitoNEET in nanodisc (To mimic function of native mitoNEET in biological membrane) by ubiquinone-10 under anerobic conditions. Our result shows that under anerobic conditions, ubiquinone-10 oxidizes the reduced [2Fe-2S] clusters of YneM-mitoNEET in nanodisc. The results indicate that ubiquinone-10 may be an intrinsic oxidant for the reduced [2Fe-2S] clusters of the native mitochondrial outer membrane protein mitoNEET (containing both membrane and soluble portion) and that native mitoNEET act as a redox enzyme that catalyzes electron transfer from FMNH2 to ubiquinone-10 in mitochondria.

Committee Chair

Huangen Ding



Included in

Biology Commons