NEWS OF THE WEEK
BACTERIAL ACID TRIPS CHEMICAL BIOLOGY: New technique reveals how pathogens endure our acidic stomachs HEN YOU GET food poisoning, the bacteria
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causing havoc in your intestines have navigated a rather treacherous journey through your acidic stomach. The molecular mechanisms by which a pathogen survives this acid environment have long kept researchers guessing. But now a team of biochemists led by Peng R. Chen of Peking University, N in China, has developed a new technique to study N the bacterial coping mechanism (Nat. Chem. Biol., O DOI: 10.1038/NChemBio.644). In addition to helping researchers better understand HN N pathogen survival, the new technique, which permits H protein-protein interactions to be studied at low pH, will likely find application in probing the biology that occurs in acidic conditions. To date, studying proteinprotein interactions at low pH has been a challenge OH because most techniques don’t work in strongly acidic O environments, Chen explains. DiZPK To study how Escherichia coli survives in the stom-
HIGH-THROUGHPUT REACTION DISCOVERY ORGANIC CHEMISTRY: Method lets chemists study thousands of transformations simultaneously
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Hartwig and Robbins discovered this coppercatalyzed alkyne hydroamination by using a highthroughput approach.
HANKS TO a new approach, chemists can now
run thousands of experiments with the hope of finding new coupling transformations, all in just a matter of days. The high-throughput method allows researchers to vary catalysts, ligands, and multiple substrates all at one time using simple laboratory equipment and mass spectrometry analysis (Science, CuCl DOI: 10.1126/ sci ence.1207922). N “It’s very simple,” says University of California, Berkeley, chemistry professor John F. Hartwig, who developed the approach with graduate student Daniel F. Robbins while the two were at the University of Illinois, Urbana-Champaign. “We hope that it’s a method that can be used by labs besides our own.” WWW.CEN-ONLINE.ORG
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ach, Chen’s group focused on the bacterium’s chaperone proteins, which prevent cellular proteins from unfolding in harmful environments. They engineered the gene for an essential chaperone protein called HdeA so that it incorporates an artificial amino acid (called DiZPK) at the site that binds client proteins. The artificial amino acid has a side chain that possesses an alkyl diazirine moiety that can cross-link to other proteins when irradiated. The team exposed this engineered E. coli to acid so HdeA would start protecting proteins from denaturation. Then they hit the bacterium with light to trap HdeA with its client proteins and used mass spectrometry to identify these clients. Among the dozens of client proteins protected by HdeA, the team detected two other chaperone proteins. This finding suggests to them that HdeA is a mother chaperone protein in a large acid-resistance network. It’s “a very clever and elegant method,” says John W. Foster, a microbiologist who studies pathogen acid resistance at the University of South Alabama. “The real value of this work is in the methodology, which could be used to probe many other chaperone-client interactions in bacteria, archaea, and eukaryotes.” Indeed, Chen says he’s currently using the approach to examine protein-protein interactions in the lysosome, the organelle in eukaryotes whose acidic interior is responsible for breaking down waste proteins and other cellular molecules.—SARAH EVERTS
To screen for new coupling reactions, the chemists took four 96-well plates and loaded each of the wells with a transition-metal-catalyst precursor, a ligand, and 17 substrates, each carrying one functional group. “We chose substrates that have about the same molecular weight so if they couple we can distinguish the products from the reactants by mass spectrometry,” Hartwig explains. To eliminate the need to run an MS analysis for each well, Hartwig and Robbins developed a method to analyze representative samples from each row and each column of a plate. They then use their MS data to pinpoint wells with products. In cases where they cannot directly deduce from the MS data which of the 17 substrates reacted, they split the substrates into four groups and rerun the reaction. They repeat this winnowing process until the reacting substrates are identified. Hartwig says he got the idea from trying to identify which portion of a Microsoft Word document was making his computer crash. Rather than go through the document page by page, he repeatedly split it in half, identifying the bad half and ignoring the rest. “The method is clever and effective without being sophisticated, and can be easily adopted by others without special equipment or expertise,” writes University of Michigan chemistry professor John Montgomery in a commentary that accompanied the piece.—BETHANY HALFORD
SEPTEMBER 12, 2011
