Electrochemically Monitoring the Acid and Acidic Urea‐Induced Unfolding of Hemoglobin and Its Electrocatalytic Ability

Abstract

Acid and acidic urea-induced unfolding of hemoglobin (Hb) was investigated using protein-film-based electrochemical method. The conformational transition of Hb was monitored through the change of direct electrochemical response of Hb. The heme groups in Hb detached from their native binding sites after pH < 4.0. Spectral experiments fully supported the results. Furthermore, the catalytic ability of Hb was enhanced 15 times under the optimal unfolded conditions of pH 2.0 PBS containing 3.0 mol L À1 urea. The method contributes to understand the relationship between function and conformation of Hb, and provides possibility of manipulating protein function by controlling its conformation.

Investigation on the Conformation Change of Hemoglobin Immobilized on MPA-modified Electrode by Electrochemical Method

“…The sudden decrease of Ipc was attributed to unfolding of the Hb, in particular changes in the heme pocket and its exposure to solvent. This was different from the research results of Zhao et al, 21 in which the current rate of sharp change was between pH 4.0 and 2.0. This was probably due to the fixed method of Hb to electrode.…”

“…Previous research 10,21 showed that the electrochemical response increased upon the unfolding of Hb induced by the denaturant urea because of the electrical active center exposure. Figure 1A shows typical cyclic voltammograms (CVs) of different chemically modified electrodes in the absence or presence of 8.0 M urea as the denaturant.…”

“…Therefore, UV-vis absorption spectrum is widely used for the research of Hb conformation change. 2,10,21 Without the presence of urea, a tight Soret band at 406 nm was obtained (Fig. 2, curve a), which was assigned to the heme monomer coordinated to His-F8 in the native state of Hb.…”

“…Because of its importance in transporting oxygen from the lungs or gills to peripheral tissues in the vascular system of animals, investigations on the structural transition of Hb are invaluable for understanding physiological events. 19,20 Acid and acidic urea-induced unfolding of Hb was investigated by Zhao et al 21 using an electrochemical method. The heme group in Hb detached from their native binding sites at pH < 4.0.…”

Haemoglobin unfolding studies at the liquid–liquid interface

“…Protein unfolding / folding mechanisms are traditionally investigated by spectroscopic techniques (fluorescence, UV-Vis, circular dichroism) [1]. However, electrochemical methods have been developed to investigate protein unfolding mechanisms using, protein filmmodified electrodes [2][3][4][5][6] and mercury electrodes [7][8]. Detection of protein unfolding at proteinfilm electrodes is generally based on the release of redox-active groups, such as the haem groups of haemoglobin (Hb).…”