Ethanol and Acetic Acid Induces Conformational Changes in Zebrafish Dihydrofolate Reductase Protein

Abstract

An equilibrium unfolding research of the recombinant zDHFR would provide information on the conformational changes of protein. In present work, the conformation of recombinant zDHFR was investigated in presence of ethanol and acetic acid. Equilibrium unfolding of recombinant zDHFR was monitored by enzymatic assay after denaturation by ethanol and acetic acid at 340 nm. Changes in secondary and tertiary structure of zDHFR with increasing ethanol and acetic acid concentrations were investigated in far-UV circular dichroism (CD) (190 to 250 nm) and fluorescence spectroscopy (emission spectra from 300 to 400 nm with an excitation wavelength of 295 nm) methods. It has been observed that in case of acetic acid-induced denaturation of zDHFR, the shift from native to denatured state occurs in a single step, whereas intermediates or non-native states are found at low concentrations of ethanol. The recent findings have significant implications for understanding how ethanol and acetic acid affect protein structure.