circumvents the thermal electron problem and makes it possible to use ECD (with the ETD modification) in widely available ion trap instruments. "We had been looking for such a reagent and hadn't found one," McLuckey says. Many natural proteins are phosphorylated, and researchers would like to know the phosphorylation sites. With conventional MS fragmentation methods, such as collisionally activated dissociation, the phosphate groups fall off, and the sites where they were located can't be determined. ETD, on the other hand, cleaves the polyamide backbone of peptide and protein cations, instead of excising side chains such as phosphate groups, making it possible to identify phosphorylation sites and making E T D applicable to the analysis of phosphorylated proteins. PEPTIDE AND PROTEIN phosphorylation mapping can also be carried out by liquid chromatography-tandem MS or by a combined chemical-enzymatic approach devised by professor of cellular and molecular pharmacology Kevan M. Shokat of the University of California, San Francisco, and coworkers [Nat. Biotechnol., 2 1 , 1047 (2003); C&EN, Sept. 1, 2003, page 31}. But such specialized techniques require extra steps apart from the usual sequencing experiments, so ETD may prove more generally useful for identifying phosphorylated peptides and proteins. "If ETD extrapolates the way one would hope," McLuckey says, "it also might be a very nice way to characterize other posttranslational modifications" in addition to phosphates, such as carbohydrate groups in glycosylated proteins, "because usually when you fragment posttranslationally modified proteins they lose the modifications," whereas with E T D they do not. Hunt and coworkers also believe that it may be possible to carry out "top-down" analyses—direct sequencing of proteins, without prior enzymatic fragmentation— by combining ETD with collisionally activated dissociation and ion-ion protontransfer techniques developed earlier by McLuckey's group. Overall, Hunt and coworkers conclude that ETD will become "an indispensable tool for peptide and protein sequence analysis in the near future and is likely to drive the development of new MS instrumentation and software." McLafferty agrees. "Having the same capabilities as ECD on a far less expensive instrument already in constant use in proteomics laboratories will be a great boon," he says. • HTTP://WWW.CEN-ONLINE.ORG
A GENETIC CODE FOR ORGANIC CHEMISTRY 'DNA display' offers new option for preparing, screening large libraries of organic compounds close proximity when the strands hybridize display makes it possible to use with each other, inducing the reagents to react in a sequence-programmed manner. short DNA sequences to direct "DTS and D N A display are the first meththe synthesis of large collecods that enable DNA to be translated into tions (libraries) of small organsynthetic molecules, including structures ic molecules. not necessarily resembling proteins," Liu The method provides a new option for explains. creating and screening libraries of small Each of the two techniques will most molecules to discover agents with desired likely prove useful for different types of synproperties for a range of research and drug discovery applications. It's like a genetic code for organic chemistry, say its developers, graduate student David R. Halpin, assistant professor of biochemistry Pehr A. B. Har- ENCODED DNA display starts with a library of bury and coworkers at Stanford DNA oligomers, one of which is shown. Each has a University [PLoS Biol., pub- pattern of coding regions (colored rectangles) that lished online June 22, http:// direct the split-pool synthesis of a small molecule dx.doi.org/10.1371/journal.p (colored balls) on the end of the oligomer. bio.0020173,-74,and-75]. theses, but both can be used to identify In DNA display, small molecules are asfunctional agents. This is done by using the sembled on the ends of DNA oligomers in techniques in iterative procedures—in a split-pool reaction network—a system of which large libraries of organic compounds reactions designed to create a large numare synthesized, active compounds from ber of diverse compounds in a small numthose libraries are identified by in vitro seber of steps. First, one constructs a library lection, and the encoding D N A that reof DNA oligomers, each containing a semains associated with selected compounds ries of coding sequences. Each coding seis then amplified for reintroduction into quence directs the DNA oligomer to one of the cycle. several possible sites in a split-pool reaction network by hybridizing with a compleA key goal is to also use the techniques mentary DNA sequence at the site. to carry out "molecular evolution," in which encoding D N A for selected compounds is A chemical subunit is then added to the also modified in an effort to create novel end of the DNA oligomer at each site, and agents with even better activity "While rea small organic compound is built up on the searchers have previously found great sucoligomer as it visits a series of sites in the cess applying evolution-based approaches network. The result is a large library of small to the discovery of proteins, RNAs, and molecules attached to the ends of the DNAs with new or improved functional oligomers that encoded their synthesis. properties," Liu says, the advent of DTS Such attachment makes iterative screenand DNA display opens up the possibility ing possible. Harbury and coworkers of evolving small-molecule organic comdemonstrated the technique by using it to pounds as well. carry out two cycles of in vitro selection (screening for binding or activity) on a liDTS and D N A display "are the types of brary of 1 million nonnatural peptides, techniques that will be needed to manage yielding a high-affinity protein ligand. the big numbers [of compounds] we hoped for in the early days of combinatorial chemDNA display has the same goal as DNAistry," comments Gerald F.Joyce, profestemplated synthesis (DTS), a technique desor of chemistry and molecular biology at veloped by David R. Liu, an associate proScripps Research Institute. They are "comfessor of chemistry and chemical biology plementary approaches that will enable at Harvard University, and coworkers. In the evolution of small organic molecules," DTS, organic reagents bound to complehesays.-STUBORMAN mentary D N A strands are brought into
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