Engineered proteases for proteomics

Potomac Affinity Proteins, Llc, North Potomac MD

Investigators

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Abstract

DESCRIPTION (provided by applicant): Proteomics is a rapidly expanding field but current methodologies remain inadequate for achieving its full potential. The most basic enzymological tool for characterizing proteins is the protease. Proteases are already an essential part of proteomic analysis but more sophisticated tools are needed to identify low abundance proteins in highly complex samples. We have engineered prototype restriction proteases which are active in denaturing conditions and which cut specifically at well-defined cognate sequence motifs. Our basic innovation would be the ability to parse a proteome into sequence-edited slices and to detect these edited portions with high resolution and high sensitivity. In Phase I we tested the prototype restriction protease for suitability in proteomic analysis and implemented a novel directed evolution methodology for the selection of proteases that cut new sequence motifs. The Phase II objective is to develop a sophisticated set of protease tools to facilitate proteomic analysis in the way restriction endonucleases have facilitated genomic analysis.