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UC Santa Cruz Study Marks Advance Toward Nanopore-Based Protein Sequencing

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Researchers at the University of California, Santa Cruz, have demonstrated an enzyme-driven method for unfolding proteins and moving them through a model α-hemolysin nanopore.

The work, detailed in a paper published this week in Nature Biotechnology, marks an advance toward nanopore-based protein sequencing, providing a proof-of-concept similar to early nanopore-based DNA sequencing work, the authors wrote.

In the study, the researchers, led by Mark Akeson, Chair of Biomolecular Engineering and Professor of Biomolecular engineering at UC Santa Cruz, used the protein unfoldase ClpX to first unfold three differentially modified Smt3 proteins and pull them through an α-HL nanopore.

"Segments of each protein could be discerned based on sequence-dependent features as the protein passed through the … transmembrane pore lumen," the researchers wrote, noting that "an optimized nanopore device could provide long reads of individual native protein strands."

When voltage is applied across a membrane containing a nanopore, molecules passing through the nanopore cause changes in conductivity that can be measured, enabling detection or sequencing of those molecules. Thus far, the technology has been applied primarily to the study of nucleic acids, with a number of commercial firms, such as Oxford Nanopore, developing it for high-throughput DNA sequencing.

Oxford Nanopore is also researching nanopore-based platforms for protein measurement. The company has yet to release any commercial protein detection devices, but in January of last year founder Hagan Bayley and other researchers from the company published a study in the Journal of the American Chemical Society on using nanopores for protein detection.

Such an approach differs from the UC Santa Cruz team, however, in that, instead of trying to pass an unfolded protein through the nanopore, Bayley and his team used the nanopore to detect the binding of target proteins to a linked aptamer.

Last year in a study published in Nature Nanotechnology a team of researchers at the Technical University of Munich used a similar approach, functionalizing nanopores with recombinant his-tagged proteins to sense target analytes (PM 3/16/2012).

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