Abstract
Dihydrofolate reductase (DHFR) has been purified over 30,000-fold from Drosophila adults with a yield of 35%, using a combination of low pH extraction, (NH4)2SO4 precipitation, Sephadex gel filtration, Affi-Gel blue affinity chromatography, ion exchange and gel filtration FPLC. The Drosophila enzyme is a soluble, 17-22 kDa monomeric protein displaying the two pH optima characteristic of eukaryotic DHFRs. The sequence of the first 23 amino acids from the amino-terminal end of the protein shows that Drosophila DHFR is more homologous to the mosquito and vertebrate DHFRs than to the prokaryotic enzymes. However, the percent similarity between the two insect enzymes is not as close as expected when compared to the virtually identical initial sequence conservation of mammalian DHFRs.
