Chronopotentiometric Analysis of Proteins at Charged Electrode Surfaces

Abstract

Protein properties and functions are strongly dependent on the structure and amino acid content. In this work, catalytic hydrogen evolution reaction (CHER) of five proteins (human serum albumin, lysozyme, β‐synuclein, H2 A and H3 histones) were studied using constant current chronopotentiometric stripping (CPS) with the aim to find out the association between protein content and its electrochemical response. We have shown that the height and potential of CPS peak H in dependence on accumulation potential differed for the studied proteins, while the CPS peak area was almost the same for all of them. CV and CPS peaks H of Cys‐containing proteins appeared at less negative potentials in comparison to proteins without Cys, suggesting easier CHER. Acidic and basic proteins not containing Cys can be also recognized due to their different CPS response after their adsorption at the positive and negative charged interface.

Chronopotentiometric Analysis of Single Histones and Histone Octamer at Charged Surfaces

“…On the other hand, the observed potential shift of peaks H was only insignificant. These results are consistent with our previous findings, [26] that decrease of height and potential shift of CPS peaks H to negative values is due to the lower ability of proteins to catalyze hydrogen evolution. By the increase of pH value, the concentration of the acidic component of the buffer decreased, as well as [H 3 O + ].…”

“…Histone H3 yielded peak H at more positive potential, E P of −1.72 V, compared to other histones. The behavior of the histone H3 was in accordance with our previous results that, cysteine‐containing peptides and proteins yield peak H at a more positive potential than those without cysteine residues [26] . Peak H of histone H4 was observed at the most negative potential, E P of −1.79 V (Figure 1A), which could be due to its greatest total value of positive charge and the highest content of arginine.…”

“…In our previous work, [26] we had shown that histone H2A and H3 behaved electrochemically differently than other mostly acidic proteins in neutral phosphate buffer, in which most of the previous experiments were performed [16,23,27] . These conditions are not ideal for histone study since there are unstable in pH lower than 7 and at low ionic strength [1] .…”

“…500 nM histones were accumulated for 180 s at a highly negative accumulation potential, E A of −1.1 V, at hanging mercury drop electrode (HMDE); subsequently, a chronopotentiogram was recorded with a stripping current, I str of −10 μA. Negative accumulation potential was chosen based on our previous results since histones accumulated at potential more positive than E A of −0.7 V did not produce well‐resolved peak H [26] . We assumed that the positively charged electroactive AA residues of histones are orienting towards negatively charged HMDE surface at the accumulation time at E A of −1.1 V. At pH 7, all studied histones yielded CPS peak H with height ranging from 15 to 40 s/V.…”

“…Better understanding the behavior of various proteins at the charged surface can subsequently lead to improving CPS analysis of protein complexes. In previous work, we described the difference between acidic and basic proteins as well as between those containing and not containing cysteine residues using CPS peak H [26] . Positively charged histones H2A and H3 behaved differently than other studied proteins, namely human serum albumin, lysozyme, and β‐synuclein, and did not yield any peak H after their adsorption at a positively charged electrode surface.…”