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MGH researchers develop faster method of engineering zinc-finger nucleases
Date:12/12/2010

A team led by Massachusetts General Hospital (MGH) researchers has developed a faster way to engineer synthetic enzymes that target specific DNA sequences for inactivation, repair or alteration. The report from the MGH Molecular Pathology Unit, being published online in Nature Methods, describes a highly effective but less labor-intensive way to generate powerful tools called zinc-finger nucleases (ZFNs).

"With our approach, called context-dependent assembly, any scientist can use either standard molecular biology techniques or commercial DNA synthesis to design ZFNs for their target gene of interest," says J. Keith Joung, MD, PhD, associate chief for Research in MGH Pathology, the study's senior author. "ZFNs are broadly applicable, powerful tools for manipulating the genomes of cells from various organisms including humans and may provide a way to efficiently correct gene mutations responsible for human disease, avoiding problems resulting from the imprecise nature of current gene therapy approaches using viral vectors."

Most human transcription factors that control whether a genetic signal is translated into a protein bind to specific DNA sequences using peptides called zinc fingers. Zinc-finger nucleases are synthetic "designer" proteins combining a zinc-finger domain, engineered to bind a particular DNA sequence, with an enzyme that breaks both DNA strands at the targeted site. While ZFNs have great potential, creating the customized proteins has been challenging.

In the simplest approach, called modular assembly, individual peptides are linked together like beads on a string to create a multi-finger protein theoretically able to recognize long DNA segments. Joung and others have shown that, in practice, modular assembly has a very low success rate for creating multi-finger proteins. This high failure rate is most likely due to "context-dependent" effects that individual zinc fingers can have on the DNA-binding
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Contact: Sue McGreevey
smcgreevey@partners.org
617-724-2764
Massachusetts General Hospital
Source:Eurekalert

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