The final steps for producing ATP from fatty acid oxidation involve the transfer of electrons into the respiratory chain, which enlists the help of many proteins and cofactors. One important enzyme is electron transfer flavoprotein (ETF), an FAD containing heterodimer, which takes electrons from several different acyl-CoA dehydrogenases that initiate fatty acid oxidation in the mitochondrial matrix and<br/>transfers them to the membrane bound electron transfer flavoprotein ubiquinone oxidoreductase. Mitochondrial conditions that create covalent alteration of amino acids in ETF can change the structure of the enzyme, which can lead to decreased functionality. Specifically, this study focused on the adduction of 4-hydroxy- 2-nonenal (4HNE), a reactive peroxylipid created by oxidative stress that can result in disease states. With the knowledge that 4HNE most readily forms adducts to the amino acids histidine, cysteine, and lysine, 4HNE was incubated with ETF followed by tryptic digestion and analyzed using high resolution liquid chromatography-mass spectrometry (LC-MS). Peptide adducted sequences of ETF identified through that process were ordered for more focused (LC-MS) study on the extent and specific location of 4HNE binding, From these studies 5 sequences on the alpha subunit and 15 sequences on the beta subunit are susceptible to 4HNE binding, with 12 sites appearing to crosslink two amino acid sequences together. The latter outcome has implications in reducing the conformational flexibility of ETF, which can limit its ability to accept electrons from its spectrum of donors.<br/>

The effect of oxidative stress on mitochondrial electron transfer flavoprotein: Pinpointing sequences adducted by 4-hydroxy-2-nonenal

255th National Meeting (2018)

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