Genetic analysis of wax ester and triacylglycerol biosynthesis in Acinetobacter calcoaceticus strain BD413

The phenomena I have investigated is the accumulation of wax esters and triacylglycerol in the form of intracellular inclusions in the gram negative aerobic bacterium Acinetobacter calcoaceticus strain BD413. This strain of A. calcoaceticus accumulates both wax esters and triacylglycerol as a means of carbon storage when it undergoes nutrient starvation.;Nutritional supplementation experiments on the wax;By expressing the identified open reading frame from A. calcoaceticus in E. coli, and assaying for the encoded protein's activity, the genes enzymatic activity was identified. The gene product was observed to catalyze the formation of fatty alcohol from acyl-CoA via a fatty aldehyde intermediate. The isolated gene has been named acr1 for acyl-CoA reductase.;A second aspect of the research described herein concerns mutants isolated following transposon mutagenesis with mini-Tn10PttKm. Flanking sequence surrounding a transposon insertion in strain 11-C7, a wax;Mutants of strain BD413 were induced using both chemical and transposon mutagenesis methods. By screening colonies for wax accumulation by staining with the lipophilic dye, Sudan black B, followed by TLC, a total of 8 transposon mutants and 21 chemically induced mutants were isolated. These mutants were separated into 3 general categories: wax...