| Aptameric Peptide for One-step Detection of Protein Kinase. | |
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MedLine Citation:
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PMID: 22533554 Owner: NLM Status: Publisher |
Abstract/OtherAbstract:
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Protein kinases are significant regulators in cell signal pathway and it is difficult to achieve quick kinase detection because traditional kinase assays normally rely on time-consuming kinase phosphorylation process. Herein, we present a novel one-step strategy to detect protein kinase by using a kinase-specific aptameric peptide-functionalized quartz crystal microbalance (QCM) electrode, in which the detection can be finished in less than 10 mins. A peptide kinase inhibitor (IP20) was used as the aptameric peptide because of its selective and strong interaction with target protein kinase (cAMP-dependent protein kinase, PKA), high stability and ease of inexpensive synthesis, presenting a new directly-recognizing element for kinase. The aptameric peptide was immobilized on the Au-coated quartz electrode through dual-thiol anchoring and the binding of his-tagged peptide with NTA/Ni(II) complex, fabricating a highly specific and stable detection platform. The interaction of aptameric peptide with kinase was monitored by QCM in real time and the concentration of protein kinase was sensitively measured by the frequency response of QCM with the low detection limit for PKA at 0.061 mU µL-1 and a linear range from 0.64 mU µL-1 to 22.33 mU µL-1. This method is rapid, reagentless and without requirement of phosphorylation process. The versatility of our aptameric peptide-based strategy has also been demonstrated by the application in kinase assay using electrochemical impedance spectroscopy. Moreover, this method was successfully applied to detect the Forskolin/3-isobutyl-1-methylxanthine (IBMX)-stimulated activation of PKA in cell lysate. |
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Authors:
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Xiahong Xu; Jiang Zhou; Xin Liu; Zhou Nie; Meng Qing; Manli Guo; Shouzhuo Yao |
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Publication Detail:
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Type: JOURNAL ARTICLE Date: 2012-4-26 |
Journal Detail:
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Title: Analytical chemistry Volume: - ISSN: 1520-6882 ISO Abbreviation: - Publication Date: 2012 Apr |
Date Detail:
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Created Date: 2012-4-26 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 0370536 Medline TA: Anal Chem Country: - |
Other Details:
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Languages: ENG Pagination: - Citation Subset: - |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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