To gain better signals in potentiometric biosensing of protein, site-selective chemical modification of amino acid residues was employed by exogenous acylation and glycation reactions of primary amines and the guanidinium group, converting them from cationic into anionic or neutral. The mass shift corresponding to the lysine and arginine adducts was confirmed only at the surface-exposed solvent-accessible residues. The site-selectivity of the charge-conversions resulted in maintained structural integrity and bioactivity of the proteins. The estimated negative charge density of bovine serum albumin (BSA) under physiological pH increased by 5-fold as a result of the formation of stable succinic lysine. Real-time measurement of protein adsorption onto the 1-undecanethiol self-assembled monolayer (SAM) on gold was detected using an extended gate-field effect transistor (FET). The potential shifts by the adsorption was 3-fold higher in succinylated BSA than origianl BSA, whereas more significant amplification of the signal (11-fold) was observed by the modifications of lysozyme. Furthermore, in situ modification of amino acids during the potentiometry was achieved for the tightly adsorbed lysozyme onto the SAM. In summary, site-selective charge-conversion provides a new type for the molecular "label" for biosensing with preserved conformational integrity of the protein.
展开▼
