Tuning the Surface Charge of Self-Assembled Monolayers on Nanoporous Gold Electrodes for Electrochemical Controlled Immobilization of Enzymes

Description

Due to their high surface-to-volume ratio and electrical conductivity as well as free-standing structure, nanoporous metals produced by electrochemical controlled etching, exhibit great potential for application as (enzymatic) biosensors. One particular interesting approach would be to use the polarizability of the electrode surface to control the immobilization of biomolecules, e.g. enzymes. To achieve this, first the metallic surface has to be modified to ensure selective and efficient binding of enzymes. For planar electrodes, the use of self-assembled monolayers (SAMs), which are organic molecules that bind spontaneously to the surface, is well established for this purpose and provide highly versatile functionalities. However, for nanoporous electrodes it is known that the self-assembling process itself and the resulting properties of the monolayer differ strongly from planar surfaces but only few studies are available. 1
In this study, nanoporous gold (npAu) is modified with 16-mercaptohexadecanoic acid (MHDA) and 2-mercaptoethanesulfonic acid (MESA) and the proton transfer reaction was investigated with cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) in phosphate buffer solution at a pH of 7.5. For MHDA, the protonation/deprotonation reaction was qualitatively well in line with the literature for planar gold electrodes, while the fraction of electrochemical controllable SAMs increased by a factor of 10 compared to a planar system. This could be attributed to the surface roughness and hence different ordering of the molecules.2 For MESA, the proton transfer reaction was observable as well, under the same conditions indicating a significant shift in pKa compared to dilute sulfonic acid.
In a second step, these findings were used to investigate the immobilization of the hydroxylase P450 BM3 (CYP102A1) containing the positively charged binding module zBasic2.3 In total, a surface coverage of 0.6 mg/m2 (equivalent to approx. 2 monolayers) was achieved. For investigation of the enzyme’s activity, the O2 consumption was monitored during the hydroxylation of lauric acid yielding an efficiency factor of 30%. The process was monitored with EIS and analyzed by means of electrical equivalent circuit providing in-situ information on the binding process. Finally, in the presentation the influence of the surface charge of the SAMs on the immobilization kinetics will be discussed

Period	5 Nov 2021
Event title	10th International Workshop on Surface Modification for Chemical and Biochemical Sensing
Event type	Conference
Location	Warsaw, PolandShow on map
Degree of Recognition	International