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Carboxyphenyl Covalent Immobilization of Heme Proteins and its Favorable Biocompatible Electrochemical and Electrocatalytic Characteristics
Author(s) -
Liu Meichuan,
Qi Yuan,
Zhao Guohua
Publication year - 2008
Publication title -
electroanalysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 128
eISSN - 1521-4109
pISSN - 1040-0397
DOI - 10.1002/elan.200704114
Subject(s) - covalent bond , chemistry , electrochemistry , amperometry , hemeprotein , redox , electron transfer , heme , cytochrome c , electrode , biocompatibility , biosensor , nuclear chemistry , inorganic chemistry , combinatorial chemistry , organic chemistry , enzyme , biochemistry , mitochondrion
A novel carboxyphenyl covalent immobilization technique has been successfully developed to realize the effective attachment of two typical heme proteins, hemoglobin (Hb) and cytochrome c (Cyt‐c), onto underlying glassy carbon electrode (GCE). Primarily, the GCE surface is functionalized with aromatic 4‐carboxyphenyl (4‐CP) group by the electrochemical reduction of diazonium cations, producing covalently linked carboxyl‐terminated active GCE surface to work as a ‘bridge’. Then, Hb and Cyt‐c are readily attached to GCE through the ‘bridge’ by functional covalently combination between NH 2 terminal groups of proteins and COOH terminal groups of 4‐CP, obtaining Hb/4‐CP/GCE and Cyt‐c/4‐CP/GCE. On both electrodes, well‐defined peaks attributing to the Fe III /Fe II couple of heme group of Hb and Cyt‐c are clearly observed with the electron transfer rate constant ( k s ) evaluated to be 2.48±0.05 s −1 and 2.73±0.05 s −1 , respectively. It is attractive that the formal potential ( E °') of the immobilized Hb and Cyt‐c are estimated to be 50 and 100 mV (vs. SCE), respectively, which are closer to the standard redox potential of native Hb and Cyt‐c in solution, owing to the good biocompatibility of 4‐CP groups. The electrodes also exhibit fast response, high sensitivity and well stability for the amperometric detection of H 2 O 2 at a fairly mild potential of 0 V without any mediators, obtaining rather small apparent Michaelis‐Menten constant ( K M app ) values of 113 μM for Hb/4‐CP/GCE and 101 μM for Cyt‐c/4‐CP/GCE. All the experimental results indicated that the covalent graft 4‐carboxyphenyl group plays an important role in constructing a “biocompatible bridge” to help the direct electron transfer of Hb and Cyt‐c with favorable biocompatibility and good bio‐ electrocatalytic affinity in virtue of the substituted aryl group only consisting of C, H and O elements, which is similar with the constitutes of organics. It makes the system of functionalized covalent immobilization of proteins onto carbon electrode a promising platform for fabricating the third‐generation biosensors. A new approach for realizing direct electrochemistry of proteins, as well as design of novel bioelectronic devices has been accordingly provided.