Nanoscale Mapping and Affinity Constant Measurement of Signal-Transducing Proteins by Atomic Force Microscopy
SCIE
SCOPUS
- Title
- Nanoscale Mapping and Affinity Constant Measurement of Signal-Transducing Proteins by Atomic Force Microscopy
- Authors
- Kim, IH; Lee, MN; Ryu, SH; Park, JW
- Date Issued
- 2011-03-01
- Publisher
- AMER CHEMICAL SOC
- Abstract
- Atomic force microscope (AFM) was used to measure the interaction force between two signal-transducing proteins, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and Ras homologue enriched in brain (Rheb), and to analyze the binding of glyceraldehyde-3-phosphate (Gly-3-P) to GAPDH. To enhance the recognition efficiency and avoid undesirable multiple interactions, the AFM probe and the substrate were each modified with a dendron, glutathione S-transferase (GST)-fused proteins were employed, and reduced glutathione (GSH) was conjugated at the apex of each immobilized dendron. The resulting median specific force between GAPDH and Rheb was 38 +/- 1 pN at a loading rate of 3.7 x 10(3) pN/s. The measurements showed that the GAPDH Rheb interaction was inhibited by binding of Gly-3-P. An adhesion force map showed individual GADPHs on the surface and that the number density of GAPDH decreased with the concentration of Gly-3-P. Maps obtained in the presence of various Gly-3-P concentrations provided information on the binding behavior, yielding a thermodynamic association constant of 2.7 x 10(5) M-1.
- Keywords
- TSC2 GAP ACTIVITY; GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE; BINDING; RHEB; SPECTROSCOPY; MTOR; RECEPTORS; EVENTS; GTPASE; KINASE
- URI
- https://oasis.postech.ac.kr/handle/2014.oak/24970
- DOI
- 10.1021/AC102695E
- ISSN
- 0003-2700
- Article Type
- Article
- Citation
- ANALYTICAL CHEMISTRY, vol. 83, no. 5, page. 1500 - 1503, 2011-03-01
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