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Session 6, Abstract 33


Garth Kong* (Ryan Mehl), Oregon State University

Oxidative stress (OS) can be characterized as heightened level of oxidants such as hydrogen peroxide, (H2O2 ) superoxide (O2.- ), and peroxynitrite (ONOO-). An increase in oxidants can also modify proteins for change in cell signaling during OS, and one example of this is the formation of 3-nitro-tyrosine (nitroTyr) on proteins. Nitrotyrosine has been observed in over 80 diseases including ALS, Alzheimer’s, atherosclerosis, Parkinson’s, ischemic brain injury due to stroke, etc. Tyrosine nitration of proteins may be part of a signaling mechanism during oxidative stress because it has been shown to be upregulated in diseased states, is site-specific to certain proteins, and reversible. However, the true function of nitrotyrosine is still debatable. Calmodulin (CaM) is a 16 kDa calcium-sensing second messenger protein that is highly conserved in eukaryote and archaea, and almost ubiquitously expressed in all cell types. It is the essential calcium signaling protein in the cell, and has been known to bind to thousands of other proteins to regulate vital functions of the cell including nitric oxide synthesis, translation regulation, apoptosis, and nonspecific phosphorylation; calmodulin is susceptible to tyrosine nitration at sites 99 and 138, although the biochemical function of tyrosine nitration of calmodulin is unclear because generating pure, site-specifically nitrated protein was not previously possible. In this study, we utilized genetic code expansion techniques to site-specifically replace natural tyrosine residues with the unnatural amino acids nitrotyrosine into calmodulin. As the hub of calcium signaling in the cell, we wanted to answer the question of how will nitrotyrosine affect the interactome of calmodulin.