Results and discussion
Folate can be efficiently conjugated to CAT and SOD
Table 1.
Folate can be effectively conjugated to CAT and SOD
CatalaseSuperoxide dismutase
Total number of lysines11220
Folates conjugated with lysines8419
The total number of lysine residues was determined based on four subunits of CAT and two subunits of SOD.
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Scheme 1.
Folate-conjugated antioxidant enzymes are endocytosed by activated macrophages and scavenge ROS produced by macrophages. Folate is conjugated to CAT using NHS/EDC chemistry. Folate-conjugated enzymes are endocytosed by activated macrophages via receptor mediated-endocytosis. The enzymes scavenge ROS in the endosome or VX765 of the macrophages, leading to reduced inflammation.
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Fig. 1.
Folate-conjugated CAT and SOD retain their enzymatic activity. (A) The effect of folate conjugation on CAT activity. CAT activity was determined by measuring the reduction rate of H2O2-free CAT (broken lines) and folate conjugated enzyme (solid lines), 3 μg/mL (gray lines) and 16 μg/mL (black lines). (B) The effect of the folate conjugation on SOD activity. SOD activity was determined by measuring the inhibition of WST-1 (2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt) oxidation—negative controls (gray line), free SOD (black broken line), and folate-conjugated SOD (black solid line).
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