Nature Methods

A new technique, presented in the August issue of Nature Methods, takes the guesswork out of protein mapping, and enables scientists to quickly determine where modifications are being introduced.

A map requires a certain level of detail to be truly useful-just knowing that there's a post office "midtown" doesn't do you nearly as much good as being able to see that it's located on the corner of 34th and Broadway. The same is true with proteins. Scientists have developed numerous techniques to detect proteins that have been modified in different ways. While helpful, this is not nearly as useful as finding out exactly where those modifications are on the protein-information that can reveal the functional importance of those changes.

Phosphorylation is one of the most important forms of protein modification, playing an essential role in cell signaling, division, and many other key tasks. Plucking phosphorylated proteins out from a complex mixture is relatively straightforward, but can be inefficient. Ruedi Aebersold and his colleagues have now upped the ante. They use specialized, soluble polymer molecules known as dendrimers as the key component in a chemical purification procedure that is simpler and much more effective, and enables the direct identification of phosphorylated amino acids.

The investigators sucessfully use their technique to characterize the total phosphorylation sites for a variety of individual signaling proteins, and find numerous previously unknown sites. In an accompanying News & Views, Eric Peters hails this work as a powerful integration of chemistry and proteomics, and an approach that may prove very useful in the investigation of a variety of other biochemical questions.

CONTACT
Ruedi Aebersold (University of Zürich, Zürich, Switzerland)
Tel: +41 1 633 3170; Email: aebersold@ismsb.biol.ethz.ch

Eric C. Peters (Genomics Institute of the Novartis Research Foundation, San Diego, CA, USA)
Tel: +1 858 812 1548; Email: epeters@gnf.org

Nature Methods

A new purification method offers scientists an efficient and budget-friendly tool for large scale protein production, without the need for pricey equipment or fancy resins, as reported in an article published online by Nature Methods.

There are hundreds of ways to isolate recombinant proteins, many of which reliably yield large amounts of pure, fully functional product. Unfortunately, the most effective systems require the use of highly specialized chromatography reagents-for example, resins that have been chemically modified to assist in separation-making large-scale protein preparation an expensive prospect.

Now, David Wood and colleagues offer a remarkably simple alternative for the efficient preparation of fully functional proteins. The process relies on a polypeptide tag with keen sensitivity to temperature and salt levels; by modulating these conditions, the tag can be reversibly rendered either soluble or insoluble, allowing simple separation of the tagged protein from other cellular proteins by centrifugation. This tag is attached to the target protein by a self-cleaving linker, and once the tagged protein has been separated in initial rounds of centrifugation, the tag can be prompted to detach itself, leaving behind only the protein of interest.

The investigators test their technique with a wide range of proteins, and demonstrate that their process consistently generates highly purified proteins that retain biological function. According to the authors, this flexible and cost-effective system could potentially be employed for virtually any protein, and offers a level of simplicity that could benefit both small labs and large-scale proteomic research efforts.

CONTACT
David W. Wood, co-author (Princeton University, Princeton, NJ, USA)
Tel: +1 609 258 5721; Email: dwood@princeton.edu

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