Development of a peptide inhibitor-based cantilever sensor assay for cyclic adenosine monophosphate-dependent protein kinase

Hyuk Sung Kwon, Ki Cheol Han, Kyo Seon Hwang, Jeong Hoon Lee, Tae Song Kim, Dae Sung Yoon, Eun Gyeong Yang

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

A highly sensitive nanomechanical cantilever sensor assay based on an electrical measurement has been developed for detecting activated cyclic adenosine monophosphate (cyclic AMP)-dependent protein kinase (PKA). Employing a peptide derived from the heat-stable protein kinase inhibitor (PKI), a magnetic bead system was first selected as a vehicle to immobilize the PKI-(5-24) peptide for capturing PKA catalytic subunit and the activity assay was applied for indirectly assessing the binding. Synergistic interactions of adenosine triphosphate (ATP) and the peptide inhibitor with the kinase were then investigated by a solution phase capillary electrophoretic assay, and by surface plasmon resonance technology which involved immobilization of the peptide inhibitor. After systemically evaluated by a homogeneous direct binding assay, the ATP-dependent recognition of the catalytic subunit of PKA by PKI-(5-24) was successfully transferred on to the nanomechanical cantilevers at protein concentrations of 6.6 pM-66 nM, exhibiting much higher sensitivity and wider dynamic range than the conventional activity assay. Thus, direct assessment of activated kinases using the cantilever sensor system functionalized with specific peptide inhibitors holds great promise in analytical applications and clinical medicine.

Original languageEnglish
Pages (from-to)344-349
Number of pages6
JournalAnalytica Chimica Acta
Volume585
Issue number2
DOIs
Publication statusPublished - 2007 Mar 7
Externally publishedYes

Fingerprint

Cyclic AMP-Dependent Protein Kinases
peptide
inhibitor
Assays
assay
sensor
Peptides
protein
Sensors
Protein Kinases
Catalytic Domain
Phosphotransferases
Adenosine Triphosphate
Surface Plasmon Resonance
Clinical Medicine
Protein Kinase Inhibitors
Immobilization
Surface plasmon resonance
Hot Temperature
Medicine

Keywords

  • Capillary electrophoresis
  • Cyclic adenosine monophosphate-dependent protein kinase
  • Fluorescence polarization
  • Heat-stable protein kinase inhibitor
  • Nanomechanical cantilever
  • Peptide inhibitor
  • Surface plasmon resonance

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Environmental Chemistry
  • Spectroscopy

Cite this

Development of a peptide inhibitor-based cantilever sensor assay for cyclic adenosine monophosphate-dependent protein kinase. / Kwon, Hyuk Sung; Han, Ki Cheol; Hwang, Kyo Seon; Lee, Jeong Hoon; Kim, Tae Song; Yoon, Dae Sung; Yang, Eun Gyeong.

In: Analytica Chimica Acta, Vol. 585, No. 2, 07.03.2007, p. 344-349.

Research output: Contribution to journalArticle

Kwon, Hyuk Sung ; Han, Ki Cheol ; Hwang, Kyo Seon ; Lee, Jeong Hoon ; Kim, Tae Song ; Yoon, Dae Sung ; Yang, Eun Gyeong. / Development of a peptide inhibitor-based cantilever sensor assay for cyclic adenosine monophosphate-dependent protein kinase. In: Analytica Chimica Acta. 2007 ; Vol. 585, No. 2. pp. 344-349.
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