Download or read book Mechanism of Protein Acetylation in Mitochondria written by Josue Baeza and published by . This book was released on 2017 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Reversible N-[epsilon]-acetylation has emerged as a widespread and regulatory modification throughout the cell. First identified on N-terminal histone tails over 50 years ago, acetylation has now been identified on over 36,000 lysine sites on organisms ranging from bacteria to mammals, highlighting the strong, conserved role for this modification. Acetylation is controlled by the opposing action of lysine acetyltransferases and deacetylases and recent evidence supports a role for nonenzymatic acetylation. Lysine acetylation influences many biochemical processes including: protein-protein and protein-DNA interactions, stability, cellular localization and enzymatic activity, which is thought to arise from neutralization of the positive charge on lysine residues at neutral pH. In mitochondria, acetylation generally acts as an inhibitory modification, altering protein interactions, lowered enzymatic activity and mislocalization of mitochondrial proteins. In this case, acetylation acts as a rheostat, modulating the degree of a biochemical process. Therefore, knowing the fraction of the protein that is modified, or stoichiometry, would provide critical information for understanding the regulatory effects of lysine acetylation. The mechanism of mitochondrial protein acetylation remains unknown. While there is a wealth of data supporting a dynamic and regulatory role for acetylation within mitochondria, there is very little evidence for an enzyme-catalyzed reaction. However, the conditions of the mitochondrial matrix: elevated acetyl-CoA levels and alkaline pH, raise the possibility that lysine acetylation can occur nonenzymatically. Throughout my thesis research, I have utilized chemical, biochemical, proteomic, and cellular studies to characterize protein acetylation. Specifically, I developed a mass spectrometry based approach to directly quantify lysine acetylation stoichiometry at the proteome-wide scale. Using this newly developed method, I measured the second order rate constants of nonenzymatic lysine acetylation using mitochondrial and non-mitochondrial proteins, highlighting the wide range of reactivities. The characterization of nonenzymatic rates of native protein demonstrates that the uncatalyzed reaction can account for the levels of acetylation in vivo. I also provide evidence that the majority of mitochondrial acetylation occurs within the mitochondria. Chapter 1 introduces lysine acetylation, the various processes controlling cellular levels including enzymatic and nonenzymatic, and why quantifying acetylation stoichiometry is needed. Chapter 2 discusses the mass spectrometry based method used for quantifying stoichiometry and its application in the model system, Escherichia coli. In chapter 3, I quantify the rates of nonenzymatic acetylation using a panel of mitochondrial and nonmitochondrial proteins, which highlights, for the first time, the wide range of lysine reactivities on native proteins. I then utilize an optimized version of the stoichiometry method to query mammalian lysine acetylation stoichiometry, revealing the wide distribution of acetylation across the cell. Conclusions, ongoing work, and future directions are presented in Chapter 5.