title: Sensing Enzymatic Activity by Exposure and Selection of DNA-Encoded Probes.
abstract: A sensing approach is applied to encode quantitative enzymatic activity information into DNA sequence populations. The method utilizes DNA-linked peptide substrates as activity probes. Signal detection involves chemical manipulation of a probe population downstream of sample exposure and application of purifying, selective pressure for enzyme products. Selection-induced changes in DNA abundance indicate sample activity. The detection of protein kinase, protease, and farnesyltransferase activities is demonstrated. The assays were employed to measure enzyme inhibition by small molecules and activity in cell lysates using parallel DNA sequencing or quantitative PCR. This strategy will allow the extensive infrastructure for genetic analysis to be applied to proteomic assays, which has a number of advantages in throughput, sensitivity, and sample multiplexing.
authors.affiliation: Department of Medicinal Chemistry and Molecular Pharmacology, Purdue Center for Cancer Research, Purdue University, 575 Stadium Mall Drive, West Lafayette, IN, 47905, USA.
authors.forename: Rachael R
authors.initials: RR
authors.lastname: Jetson
authors.affiliation: Department of Medicinal Chemistry and Molecular Pharmacology, Purdue Center for Cancer Research, Purdue University, 575 Stadium Mall Drive, West Lafayette, IN, 47905, USA. cjk@purdue.edu.
authors.forename: Casey J
authors.identifier: 0000-0003-2964-3520
authors.initials: CJ
authors.lastname: Krusemark
mesh_terms: D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000235:genetics*; D015342:DNA Probes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation; D051231:Farnesyltranstransferase / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D010447:Peptide Hydrolases / Q000032:analysis / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011494:Protein Kinases / Q000032:analysis / Q000378:metabolism*
chemical_list: D015342:DNA Probes; D004247:DNA; D051231:Farnesyltranstransferase; D011494:Protein Kinases; D010447:Peptide Hydrolases
keywords: DNA; analytical methods; biosensors; enzyme assays; polymerase chain reaction