Effective electrochemical method for investigation of hemoglobin unfolding based on the redox property of heme groups at glassy carbon electrodes.
This study demonstrates a facile and effective electrochemical method for investigation of hemoglobin (Hb) unfolding based on the electrochemical redox property of heme groups in Hb at bare glassy carbon (GC) electrodes. In the native state, the heme groups are deeply buried in the hydrophobic pockets of Hb with a five-coordinate high-spin complex and thus show a poor electrochemical property at bare GC electrodes. Upon the unfolding of Hb induced by the denaturant of guanidine hydrochloride (GdnHCl), the fifth coordinative bond between the heme groups and the residue of the polypeptides (His-F8) is broken, and as a result, the heme groups initially buried deeply in the hydrophobic pockets dissociate from the polypeptide chains and are reduced electrochemically at GC electrodes, which can be used to probe the unfolding of Hb. The results on the GdnHCl-induced Hb unfolding obtained with the electrochemical method described here well coincide with those studied with other methods, such as UV-vis spectroscopy, fluorescence, and circular dichroism. The application of the as-established electrochemical method is illustrated to study the kinetics of GdnHCl-induced Hb unfolding, the GdnHCl-induced unfolding of another kind of hemoprotein, catalase, and the pH-induced Hb unfolding/refolding.
Li
Xianchan
X
Beijing National Laboratory for Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China.
Zheng
Wei
W
Zhang
Limin
L
Yu
Ping
P
Lin
Yuqing
Y
Su
Lei
L
Mao
Lanqun
L
D000818:Animals; D002244:Carbon / Q000737:chemistry*; D002374:Catalase / Q000737:chemistry; D020134:Catalytic Domain; D002417:Cattle; D004563:Electrochemistry; D004566:Electrodes; D005898:Glass / Q000737:chemistry*; D019791:Guanidine / Q000494:pharmacology; D006418:Heme / Q000378:metabolism*; D006454:Hemoglobins / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D010084:Oxidation-Reduction; D010455:Peptides / Q000378:metabolism; D011489:Protein Denaturation / Q000187:drug effects; D020673:Protein Renaturation; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D013057:Spectrum Analysis
D006454:Hemoglobins; D010455:Peptides; D021122:Protein Subunits; D006418:Heme; D002244:Carbon; D002374:Catalase; D019791:Guanidine