Kinase Sensing Based on Protein Interactions at the Catalytic Site. Issue 17 (9th February 2022)

• Record Type: Journal Article
• Authors: Solomon, Ohad; Sapir, Hannah; Mervinetsky, Evgeniy; Chen, Yu‐Ju; Friedler, Assaf; Yitzchaik, Shlomo
• Is Part Of: Chemistry. Volume 28:Issue 17(2022)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2022-02-09
• Subjects: biosensing -- electrochemical biosensing -- kinase sensing -- protein-protein interactions -- surface chemistry; Chemistry -- Periodicals; 540
• DOI: 10.1002/chem.202104227
• Languages: English
• ISSNs: 0947-6539

Abstract

Abstract: The role kinases play in regulating cellular processes makes them potential biomarkers for detecting the onset and prognosis of various diseases, including many types of cancer. Current kinase biosensors, including electrochemical and radiometric methods, rely on sensing the ATP‐dependant enzymatic phosphorylation reaction. Here we introduce a new type of interaction‐based electrochemical kinase biosensor that does not require any chemical labelling or modification. The basis for sensing is the interactions between the catalytic site of the kinase and the phosphorylation site of its substrate rather than the phosphorylation reaction. We demonstrated this concept with the ERK2 kinase and its substrate protein HDGF, which is involved in lung cancer. A peptide monolayer derived from the HDGF phosphorylation site was adsorbed onto a gold electrode and was used to sense ERK2 without ATP. The sensitivity of the assay was down to 10 nM of ERK2, corresponding with the range of its cellular concentrations. Surface chemistry analysis confirmed that ERK2 was bound to the HDGF peptide monolayer. This increased the permeability of redox‐active species through the monolayer and resulted in ERK2 electrochemical sensing. Since our detection approach is based on protein‐protein interactions and not on the enzymatic reaction, it can be further utilized for more selective detection of different types of enzymes. Abstract : An electrochemical biosensor, based on the interaction between the catalytic site of the kinase and the phosphorylation site of its substrate for detection, is described. The phosphorylation site peptide of HDGF, a substrate of ERK2, was absorbed on gold electrodes to electrochemically sense ERK2 without ATP. Surface chemistry analysis showed that ERK2 was bound to the peptide monolayer, which changed the packing of the layer, resulting in increased permeability of redox‐active species through the layer and electrochemical sensing.