NEWS OF THE WEEK
EVOLVED ENZYME ADDS SUGARS CARBOHYDRATES: Mutant
glycosyltransferase makes range of aryl glycosides for drug discovery
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LYCOSYLTRANSFERASE ENZYMES area pretty fussy lot. They catalyze the addition of sugars to other sugars or to natural products, but they'll only do this for a limited range of substrates. A research team has now created a modified glycosyltransferase that is considerably more liberal, catalyzing the addition of a variety of "donor" sugars to a wide range of aryl "acceptor" substrates to yield a range of aryl glycosides. Sugar-containing compounds tend to be good drug candidates, and the mutant glycosyltransferase will make it easier to synthesize sugar-derivatized compounds that can be tested for bioactivity. The modified glycosyltransferase was developed by postdocs Gavin J. Williams and Changsheng Zhang and pharmaceutical sciences professor Jon S. Thorson of the University ofWisconsin, Madison. They took a bacterial glycosyltransferase that works with a limited range of donors and acceptors and turned it into one that's much more "promiscuous" (Nat Chem. Biol, DOI: io.i038/nchembio.2007.28). The natural glycosyltransferase they started with accelerates the addition of either ofjust two sugars, glucose or deoxyglucose, to the natural product oleandomycin in Streptomyces bacteria. Using a technique called directed evolution, the researchers generated random mutations in the gene for the enzyme, introduced the mutated genes into bacteria, and used
them to express mutant enzymes. They screened the mutants for activity by using engineered fluorescent acceptor substrates that stopped shining when enzymes attached sugars to them. This strategy enabled them to identify several mutants with greatly liberalized substrate selectivity. They combined the most promising mutations in one gene to create their promiscuous glycosyltransferase. Whereas the natural enzyme works with just two sugar donors, the mutant version used 15 of 22 the researchers tried. And instead of adding the sugars only to oleandomycin, it adds them to a wide range of aryl organic compounds. Thorson's group is currently studying the enzyme to see whether it works on other substrates as well. Thorson notes that the only other glycosyltransferase that has been "evolved" before attaches sugars to other sugars. In contrast, the one evolved by Thorson's group adds sugars to aryl compounds, a type of activity likely to be more broadly useful for drug discovery. "Our understanding of structure and specificity is not yet sufficient for this to be done on a design basis, hence the need for directed-evolution strategies," comments chemistry professor Stephen G. Withers of the University of British Columbia, who evolved the earlier glycosyltransferase. He says the Thorson group's enzyme "will be of particular importance for the generation of antibiotics that contain unusual sugars."—STU BORMAN
Mutant glycosyltransferase (left) catalyzes addition of a variety ofsugars (red) to aryl organics bearing a range ofsubstituents (R') and to oleandomycin (lower right), the native enzyme's substrate. Mutated residues are spheres, and substrates are stick representations.
BIOTECHNOLOGY Isis and Alnylam joint venture will capitalize on microRNA expertise With the launch of an independent joint venture called Regulus Therapeutics, Isis Pharmaceuticals and Alnylam Pharmaceuticals hope to become a force in the emerging field of therapeutics that target microRNAs. Both firms will contribute intellectual property, technology, and expertise in microRNAs to Regulus, and Alnylam will lay out $10 million in initial funding. Future funding will come equally from Isis and Alnylam. MicroRNAs are small, single-stranded RNAs that are about 20 nucleotides in length. They serve as the gatekeepers for messenger RNAs, regulating mRNAs' ability to relay the instructions for protein
production to their intended targets. The roughly 500 microRNAs that have been identified are believed to regulate expression of one-third of all human genes, according to Isis CEO Stanley Crooke. The ubiquity of microRNAs, as well as their ability to broadly modulate disease, has quickly made them an intriguing therapeutic target. "Antagonism of microRNA defines a new strategy to target multiple points in entire pathways in human disease, not just single disease targets," Crooke said during a conference call to discuss Regulus. Though it often takes decades to translate the identification of new drug
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targets into actual drugs, Isis and Alnylam believe the timeline for developing microRNA therapeutics could be significantly abbreviated. Regulus will benefit from Isis' expertise in making secondgeneration antisense compounds, which the companies say is "the ideal class of drugs" to inhibit microRNAs. The new company's most advanced drug candidate targets miR-122, a microRNA implicated in hepatitis C. So far, Regulus' competition is limited. Israelbased Rosetta Genomics and Sirna Therapeutics, which was acquired last year by Merck & Co., are also vying for a position in the burgeoning field.-LISA JARVIS
