Improved Triacylglycerol Production in Acinetobacter baylyi ADP1 by Metabolic Engineering

Abstract

BackgroundTriacylglycerols are used in various purposes including food applications, cosmetics, oleochemicals and biofuels. Currently the main sources for triacylglycerol are vegetable oils, and microbial triacylglycerol has been suggested as an alternative for these. Due to the low production rates and yields of microbial processes, the role of metabolic engineering has become more significant. As a robust model organism for genetic and metabolic studies, and for the natural capability to produce triacylglycerol, Acinetobacter baylyi ADP1 serves as an excellent organism for modelling the effects of metabolic engineering for energy molecule biosynthesis.ResultsBeneficial gene deletions regarding triacylglycerol production were screened by computational means exploiting the metabolic model of ADP1. Four deletions, acr1, poxB, dgkA, and a triacylglycerol lipase were chosen to be studied experimentally both separately and concurrently by constructing a knock-out strain (MT) with three of the deletions. Improvements in triacylglycerol production were observed: the strain MT produced 5.6 fold more triacylglycerol (mg/g cell dry weight) compared to the wild type strain, and the proportion of triacylglycerol in total lipids was increased by 8-fold.ConclusionsIn silico predictions of beneficial gene deletions were verified experimentally. The chosen single and multiple gene deletions affected beneficially the natural triacylglycerol metabolism of A. baylyi ADP1. This study demonstrates the importance of single gene deletions in triacylglycerol metabolism, and proposes Acinetobacter sp. ADP1 as a model system for bioenergetic studies regarding metabolic engineering.

Improving Fatty Acid Availability for Bio-Hydrocarbon Production in Escherichia coli by Metabolic Engineering

“…As shown in Figure 6, using M9, the most cost-effective medium for industrial operation, only a small amount of TAG, 13 mg/g dry cell weight, was produced. It appears that the minimal medium does not provide a favorable condition for TAG production, as demonstrated here and in a previous study by Santala et al [45]. The rich medium was more effective and the largest amount of TAG, 49 mg/g dry cell weight, was observed in LB supplemented with 1% glucose and 2 g/L sodium bicarbonate strain.…”

Real-Time monitoring of intracellular wax ester metabolism

“…The gene knock-out of poxB has been shown not to negatively affect the WE production or growth in A. baylyi ADP1 [11] and was thus used as a target site for gene replacement. The strain ADP1 wild type was transformed with the gene cassette and selected on LA plate containing 50 μg/ml chloramphenicol.…”

Metabolic Engineering of Acinetobacter baylyi ADP1 for Improved Growth on Gluconate and Glucose

“…HPLC was carried out using the same equipment and operation condition as those used by Santala et al (12), except for the column, which was a Rezex RHM-Monosaccharide Hϩ (8%) column (Phenomenex, Torrance, CA). Standards (0.05 to 10 mM) were prepared in sterile ion-exchanged water and were run in duplicates.…”

“…These traits make it an attractive host organism for various biotechnological applications (9). The suitability of A. baylyi has been studied, for example, in production of cyanophycin (10), emulsan (11), and storage lipids (12). A. baylyi has also been used in D-xylulose production from D-xylose, where the cells were used in oxidation of residual D-xylose to D-xylonic acid (13).…”