Kwon, Hyuk-Sung Han, Ki-Cheol Hwang, Kyo Seon Lee, Jeong Hoon Kim, Tae Song Yoon, Dae Sung Yang, Eun Gyeong 2024-01-21T01:06:30Z 2024-01-21T01:06:30Z 2021-09-04 2007-03-07 0003-2670 https://pubs.kist.re.kr/handle/201004/134542 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. (c) 2007 Elsevier B.V. All rights reserved. English ELSEVIER SCIENCE BV FLUORESCENCE POLARIZATION ASSAY CAPILLARY-ELECTROPHORESIS CATALYTIC SUBUNIT TYROSINE KINASES MICROCANTILEVER IMMUNOASSAY BINDING Development of a peptide inhibitor-based cantilever sensor assay for cyclic adenosine monophosphate-dependent protein kinase Article 10.1016/j.aca.2006.12.037 1 ANALYTICA CHIMICA ACTA, v.585, no.2, pp.344 - 349 ANALYTICA CHIMICA ACTA 585 2 344 349 scie scopus 000244328100020 2-s2.0-33846591535 Chemistry, Analytical Chemistry Article FLUORESCENCE POLARIZATION ASSAY CAPILLARY-ELECTROPHORESIS CATALYTIC SUBUNIT TYROSINE KINASES MICROCANTILEVER IMMUNOASSAY BINDING peptide inhibitor heat-stable protein kinase inhibitor cyclic adenosine ruonophosphate-dependent protein kinase nanomechanical cantilever surface plasmon resonance capillary electrophoresis fluorescence polarization