The Rhodococcus opacus TadD protein mediates triacylglycerol metabolism by regulating intracellular NADPH poolsReport as inadecuate




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Microbial Cell Factories

, 12:104

First Online: 09 November 2013Received: 02 September 2013Accepted: 05 November 2013

Abstract

BackgroundThe Gram-positive actinomycete Rhodococcus opacus is widely studied for its innate ability to store large amounts of carbon in the form of triacylglycerol TAG. Several groups have demonstrated that R. opacus PD630 is capable of storing anywhere from 50 to 76% of its cell dry weight as TAG. While numerous studies have focused on phenomenological aspects of this process, few have sought to identify the underlying molecular and biochemical mechanisms responsible for the biosynthesis and storage of this molecule.

ResultsHerein we further our previous efforts to illuminate the black box that is lipid metabolism in actinomycetes using a genetic approach. Utilizing a simple, colorimetric genetic screen, we have identified a gene, referred to herein as tadD t riacylglycerol a ccumulation d eficient, which is critical for TAG biosynthesis in R. opacus PD630. Furthermore, we demonstrate that the purified protein product of this gene is capable of oxidizing glyceraldehyde-3-phosphate, while simultaneously reducing NADP to NADPH. Supporting this biochemical data, we observed that the ratio of NADPH to NADP is elevated in wildtype cultures grown under lipid production conditions as compared to cultures grown under vegetative growth conditions, while the mutant strain demonstrated no change irrespective of growth conditions. Finally, we demonstrate that over-expressing a putative phosphorylative glyceraldehyde-3-phosphate dehydrogenase leads to decreased TAG production during growth on TAG accumulation conditions.

ConclusionTaken together, the data support the identification of a key metabolic branch point separating vegetative growth and lipid accumulation lifestyles in Rhodococcus.

KeywordsRhodococcus opacus PD630 GapN triacylglycerol Electronic supplementary materialThe online version of this article doi:10.1186-1475-2859-12-104 contains supplementary material, which is available to authorized users.

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Author: Daniel P MacEachran - Anthony J Sinskey

Source: https://link.springer.com/







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