The antioxidant activity of recombinant rat fatty acid binding protein 1 T94A variant
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Date
2017
Authors
Le, Tyson
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Abstract
Liver fatty acid binding protein (FABP1) is present at high levels in the liver and functions as an antioxidant. A single nucleotide polymorphism was found resulting in the FABP1 T94A variant. Currently, it is not known how this mutation affects the antioxidant activity of FABP1 T94A. Rat cDNA within pGEX-6p-2 vector was isolated from bacterial cells and the mutation was induced using site-directed mutagenesis. The plasmid was transformed into competent cells that were cultured and used for the expression of FABP1 T94A. The protein was purified using the GST affinity tag system and its antioxidant capacity was assessed using DCF fluorescence assay. FABP1 T94A showed that the mutation did not change the protein’s ability to scavenge hydroxyl radicals. Thiobarbituric Acid Reactive Substances Assay was used to determine the FABP1 T94A’s antioxidant activity in simulated hydrophilic and lipophilic environments induced by the azo compounds AAPH and MeO-AMVN. T94A was able to act as an antioxidant in both cases performing almost equivalent to FABP1. However, when bound with fatty acids (palmitate and alpha-bromo palmitate) the ability of T94A as an antioxidant may be affected. In conclusion, The T94A variation of FABP1 does not have a loss of function in regards to acting as an antioxidant but the extent of function may be influenced by ligand binding.
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Keywords
Liver fatty acid binding protein, FABP1, Antioxidant, T94A, Mutant variant, Oxidative stress