6533b856fe1ef96bd12b2988

RESEARCH PRODUCT

Oxidation of dobutamine and dopamine by horseradish peroxidase

subject

description

Dobutamine and dopamine have been previously shown to interfere with enzymatic diagnostic tests and different mechanisms responsible for this effect have been postulated. We have studied the oxidation by horseradish peroxidase (HRP) of dopamine, dobutamine, and its analog with the phenol group blocked by methylation. Oxidation of dobutamine was much faster than dopamine, as reported before, whereas the methylated analog of dobutamine was oxidized at intermediate rate. This demonstrates that the phenol group in dobutamine is oxidized preferentially by HRP and acts as an electron-transfer mediator in oxidation of the catechol group. Different oxidation rates of catechol groups in dopamine and the methylated analog of dobutamine can only be explained by steric and polar/hydrophobic factors, not by differences in electron densities in aromatic rings, as postulated before for alkylcatechols. Their Km values are primarily responsible for this difference. Molecular docking of dobutamine and its methylated analog to the active site of HRP showed that for both compounds orientations with the catechol group directed towards the heme were slightly favored. However, the distance between the heme iron and the 4-OH group in these orientations was bigger than the distance between the iron ion and the oxygen atom of the phenol/methoxyphenyl groups, when they were placed in the active site. This explains preferential oxidation of the phenol group in dobutamine by HRP. Distance of the oxidizable group (phenol or catechol) to the heme cofactor was determined by interaction of the functional groups in the side chain acting as hydrogen bond donors (amino or hydroxyl group) with the peptide backbone of the Ala136-Pro139 region near the entrance to the active site.