U-M scientists develop tool to probe role of oxidative stress in aging, disease

HD Lighthouse Contributing Editor's Comment: Oxidative stress plays a role in aging and neurodegenerative diseases like Huntington's Disease.

-- Marsha L. Miller, Ph.D.
Posted to the HDL: 02-16-2008

Ursula Yakob, Associate Professor

Quantifying changes in the thiol redox proteome upon oxidative stress in vivo

Lars I. Leichert, Florian Gehrke, Harini V. Gudiseva, Tom Blackwell, Marianne Ilbert, Angela K. Walker, John R. Strahler, Philip C. Andrews, and Ursula Jakob

the press release

ANN ARBOR, Mich.---Oxygen, although essential for human life, can turn into an aggressive chemical that is outright toxic to important molecules inside our cells. This "oxidative stress" is associated with many diseases, such as Alzheimer's, heart disease and cancer, and has been suggested to be the culprit underlying aging.

In an article published online Feb. 14 in the journal Proceedings of the National Academy of Sciences (PNAS), University of Michigan researchers led by associate professor Ursula Jakob report on a new method that allows them to observe how oxidative stress affects the major building blocks of a cell, the proteins. The new technique, called OxICAT, makes it possible to quantify the oxidation state of thousands of different proteins in a single experiment.

Jakob was intrigued to find many proteins that are not permanently damaged by reactive oxygen species but actually use amino acids known as cysteines to sense oxidative stress.

"In my lab, we have been working for a long time on proteins that use cysteine as a reactive oxygen sensor," Jakob said. "With this new technique, we discovered scores of novel proteins that are sensitive towards reactive oxygen species. Interestingly, we found that many of the proteins that we identified are important for the cells to survive oxidative stress conditions." Jakob and her team now are using this powerful technique to gain fundamental insights into the molecular mechanism of aging and the role that oxidative stress plays in this process.

"Because oxidative stress plays such a prominent role in all these diseases, we want to understand why some cells and organisms can cope with the dangers of oxidative stress, while others die," said Lars Leichert, a postdoctoral research fellow in Jakob's lab and first author of the study. Such insights might lead to the development of more powerful and effective anti-oxidant strategies.

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The PNAS paper showcases a joint effort of scientists from U-M's Department of Cellular, Molecular and Developmental Biology, the Department of Human Genetics, the U-M based National Resource for Proteomics and Pathways (NRPP), and the Michigan Proteome Consortium (MPC), also at U-M. Philip Andrews, a professor of biological chemistry whose lab performed the mass spectrometry for the study, said the work demonstrates the importance of investment in such high technology infrastructure as the MPC (funded by the State of Michigan) and the NRPP (funded by the National Center for Research Resources) for biomedical research, leading to significant new discoveries and increased competitive advantage for researchers.

the journal article

Antimicrobial levels of reactive oxygen species (ROS) are produced by the mammalian host defense to kill invading bacteria and limit bacterial colonization. One main in vivo target of ROS is the thiol group of proteins. We have developed a quantitative thiol trapping technique termed OxICAT to identify physiologically important target proteins of hydrogen peroxide (H2O2) and hypochlorite (NaOCl) stress in vivo. OxICAT allows the precise quantification of oxidative thiol modifications in hundreds of different proteins in a single experiment. It also identifies the affected proteins and defines their redox-sensitive cysteine(s). Using this technique, we identified a group of Escherichia coli proteins with significantly (30–90%) oxidatively modified thiol groups, which appear to be specifically sensitive to either H2O2 or NaOCl stress. These results indicate that individual oxidants target distinct proteins in vivo. Conditionally essential E. coli genes encode one-third of redox-sensitive proteins, a finding that might explain the bacteriostatic effect of oxidative stress treatment. We identified a select group of redox-regulated proteins, which protect E. coli against oxidative stress conditions. These experiments illustrate that OxICAT, which can be used in a variety of different cell types and organisms, is a powerful tool to identify, quantify, and monitor oxidative thiol modifications in vivo.

Source: Proceedings of the Nat'l Academy of Sciences of the USA. February 14, 2008