Molecularly Engineered Charge-Conversion of Proteins for Sensitive Biosensing

摘要

To gain better signals in potentiometric biosensing ofnprotein, site-selective chemical modification of amino acidnresidues was employed by exogenous acylation and glycationnreactions of primary amines and the guanidiniumngroup, converting them from cationic into anionic ornneutral. The mass shift corresponding to the lysine andnarginine adducts was confirmed only at the surfaceexposednsolvent-accessible residues. The site-selectivitynof the charge-conversions resulted in maintained structuralnintegrity and bioactivity of the proteins. The estimatednnegative charge density of bovine serum albuminn(BSA) under physiological pH increased by 5-fold as anresult of the formation of stable succinic lysine. Real-timenmeasurement of protein adsorption onto the 1-undecanethiolnself-assembled monolayer (SAM) on gold wasndetected using an extended gate-field effect transistorn(FET). The potential shifts by the adsorption was 3-foldnhigher in succinylated BSA than origianl BSA, whereasnmore significant amplification of the signal (11-fold) wasnobserved by the modifications of lysozyme. Furthermore,nin situ modification of amino acids during the potentiometrynwas achieved for the tightly adsorbed lysozyme ontonthe SAM. In summary, site-selective charge-conversionnprovides a new type for the molecular “label” for biosensingnwith preserved conformational integrity of the protein.