The development of bacterial cell factories to produce fuels and chemicals requires an abundant and inexpensive carbon source to be cost-competitive. While cellulosic biomass (CB) is a suitable candidate, it has usually required costly pre-treatment processes for degradation of its constituent hemicellulose and cellulose into easily metabolizable sugars. A bacterium with the capability of directly degrading and consuming CB could minimize these pretreatments. The marine bacterium Saccharophagus degradans 2-40 is one of a select group of organisms that natively consumes cellulose and hemicellulose from several types of CB including switchgrass, corn stover and newsprint. S. degradans degrades and transports these forms of CB into simple sugars using a combination of 128 glycoside hydrolases. Interestingly, S. degradans does not transition instantaneously from glucose to CB, but requires an acclimation. This compels the investigation of metabolic changes required to utilize more complex saccharides. Toward this objective, we performed 13C-assisted metabolic flux analysis (13C MFA) on S. degradans to compare the metabolism of a simple saccharide (glucose) with that of an indicator of CB metabolism (cellobiose, a β-D [1-4] glucose dimer). We performed parallel 13C isotope labeling experiments utilizing U-13C and 1-13C sugars. Metabolomic and isotopic data revealed unique flux modes that favor NADPH production during both glucose and cellobiose metabolism. Furthermore, we observed apparent carbon diversion from glucose (10% to 20%), but not from cellobiose, to another sugar that remained in the growth media. Toward identifying a putative enzyme responsible for synthesizing this sugar and the enzyme's intracellular location, as well as to comprehensively understand the metabolic biology of this fascinating bacterium, we will report subsequent growth experiments as well as steady-state and instationary isotope labeling experiments using complementary labeling strategies.

Metabolic biology of Saccharophagus degradans, a cellulosic biomass-digesting bacterium, as revealed by 13C metabolic flux analysis

251st National Meeting (2016)

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