Scripps Research chemist devises new method to quantify protein changes

JUPITER, FL, January 5, 2011 — A scientist from the Florida campus of The Scripps Research Institute has devised a new method of analyzing and quantifying changes in proteins that result from a common chemical process. The new findings could provide new insights into the effects of a highly destructive form of stress on proteins in various disease models, particularly cancer.

The study, published January 5, 2011, in the online Early View of the journal Angewandte Chemie, was designated by the journal as a “very important paper,” a distinction bestowed on less than five percent of its publications.

“This new technique allows us to home in on which proteins are modified to a significant extent during periods of stress and how that changes during disease progression,” said Kate Carroll, an associate professor in the Scripps Research Department of Chemistry who conducted the study with Young Ho Seo, a research fellow at The University of Michigan. “It gives us the chance to look more closely at targets for possible therapeutic intervention. From a practical standpoint, the technique is simple and will be accessible to biologists and chemists alike.”

The new technique focuses on the process of cysteine S-hydroxylation, which plays a significant role in a number of events related to physiology in both health and disease, including the regulation of signaling proteins in various disease states.

The ability of the new technique to focus on signaling pathways, particularly in diseases such as cancer, is critical.

“Chronic disease states such as cancer can involve the modification of signaling proteins through S-hydroxylation, but other housekeeping proteins may also be targets,” she said. “Key to distinguishing which of these proteins may be involved in pathogenesis is the ability to measure the amount of S-hydroxylation at specific sites within a protein. Now you’ll be able to tell. This should help accelerate target identification in these disease-related signaling pathways and allow us to focus on proteins that are important to the process.”

During periods of cellular stress, caused by factors such as exposure to UV radiation or many disease states, the level of highly reactive oxygen-containing molecules can increase, resulting in inappropriate modification of proteins and cell damage.

One oxidant, hydrogen peroxide, functions as a messenger that can activate cell proliferation through oxidation of cysteine residues in signaling proteins, producing sulfenic acid (i.e., S-hydroxylation); cysteine is an amino acid is synthesized in the body.

Extending the Gains of an Earlier Study

In a 2009 study, Carroll found that sulfenic acid served as an early warning biomarker of the reaction between hydrogen peroxide and cysteine. Carroll tagged the miniscule reaction target with a fluorescent dye antibody. With it, Carroll was able to read the levels of sulfenic acid levels in various cell lines, including breast cancer cells.

The new technique takes those findings several steps further by allowing scientists not only to quantify the modifications to various proteins, but also to monitor these changes at the level of individual cysteines within a single protein.

Carroll used a class of reagents called isotope-coded dimedone and iododimedone, which traps and tags sulfenic acids, allowing the cysteine sites and modified proteins to be easily identified. These probes, which are highly selective for sulfenic acid, allow the S-hydroxylation process to be monitored at the exact site of the modification.

The tagged proteins can be then be analyzed by mass spectrometry, a standard technology used to determine the precise make-up of proteins and other molecules.

“This technique should be widely accessible to the scientific community because it’s so simple,” Carroll said. “It should allow researchers to identify proteins with altered S-hydroxylation profiles whose function may lend insight into events in disease progression and have utility as potential markers for disease detection.”

The study, “Quantification of Protein Sulfenic Acid Modifications Using Isotope-Coded Dimedone and Iododimedone (ICDID),” was funded by the Camile Henry Dreyfus Teacher Scholar Award and the American Heart Association Scientist Development Award. For more information on the paper, see http://onlinelibrary.wiley.com/doi/10.1002/anie.201007175/abstract

About The Scripps Research Institute

The Scripps Research Institute is one of the world’s largest independent, non-profit biomedical research organizations, at the forefront of basic biomedical science that seeks to comprehend the most fundamental processes of life. Scripps Research is internationally recognized for its discoveries in immunology, molecular and cellular biology, chemistry, neurosciences, autoimmune, cardiovascular, and infectious diseases, and synthetic vaccine development. An institution that evolved from the Scripps Metabolic Clinic founded by philanthropist Ellen Browning Scripps in 1924, Scripps Research currently employs approximately 3,000 scientists, postdoctoral fellows, scientific and other technicians, doctoral degree graduate students, and administrative and technical support personnel. Headquartered in La Jolla, California, the institute also includes Scripps Florida, whose researchers focus on basic biomedical science, drug discovery, and technology development. Scripps Florida is located in Jupiter, Florida. For more information, see www.scripps.edu .

1 COMMENT

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