Complete genome sequence, metabolic model construction and phenotypic characterization of Geobacillus LC300, an extremely thermophilic, fast growing, xylose-utilizing bacterium

Abstract

We have isolated a new extremely thermophilic fast-growing Geobacillus strain that can efficiently utilize xylose, glucose, mannose and galactose for cell growth. When grown aerobically at 72 °C, Geobacillus LC300 has a growth rate of 2.15 h−1 on glucose and 1.52 h−1 on xylose (doubling time less than 30 minutes). The corresponding specific glucose and xylose utilization rates are 5.55 g/g/h and 5.24 g/g/h, respectively. As such, Geobacillus LC300 grows 3-times faster than E. coli on glucose and xylose, and has a specific xylose utilization rate that is 3-times higher than the best metabolically engineered organism to date. To gain more insight into the metabolism of Geobacillus LC300 its genome was sequenced using PacBio’s RS II single-molecule real-time (SMRT) sequencing platform and annotated using the RAST server. Based on the genome annotation and the measured biomass composition a core metabolic network model was constructed. To further demonstrate the biotechnological potential of this organism, Geobacillus LC300 was grown to high cell-densities in a fed-batch culture, where cells maintained a high xylose utilization rate under low dissolved oxygen concentrations. All of these characteristics make Geobacillus LC300 an attractive host for future metabolic engineering and biotechnology applications.

Metabolism of the fast-growing bacterium Vibrio natriegens elucidated by 13C metabolic flux analysis

“…The results of this analysis are provided in Supplemental Materials and are shown in Fig. 3, where the biomass composition of V. natriegens is compared to the reported composition of E. coli (Long and Antoniewicz, 2014b) and the fast-growing thermophile Geobacillus LC300 (Cordova et al, 2015; Cordova and Antoniewicz, 2016). Proteins were the most abundant component of V. natriegens biomass (47% of dry weight), followed by RNA (29%), lipids (7%) and glycogen (3%).…”

“…growth rate, glucose uptake rate, and oxygen uptake rate) of V. natriegens to three other strains (Fig 6): wild-type E. coli (Leighty and Antoniewicz, 2013), adaptively evolved E. coli (Sandberg et al, 2016; Long et al, 2017), and the fast-growing extreme thermophile Geobacillus LC300 (Cordova et al, 2015; Cordova and Antoniewicz, 2016). Like V. natriegens , the E. coli characteristics reflect growth at 37 °C, while those of Geobacillus LC300 are from growth at 72 °C.…”

Fast growth phenotype of E. coli K-12 from adaptive laboratory evolution does not require intracellular flux rewiring

“…Identifying a growth-optimal proteome and regulatory mechanisms by which it can be achieved will be useful in engineering efficient strains. Understanding the hard constraints of E. coli performance, which in addition to the proteome can also include membrane space limitations (Liu et al, 2014), may motivate the development of alternative high-performance organisms for future applications (Cordova et al, 2015; Lee et al, 2016). …”

13C metabolic flux analysis of three divergent extremely thermophilic bacteria: Geobacillus sp. LC300, Thermus thermophilus HB8, and Rhodothermus marinus DSM 4252

“…Key advantages of thermophiles include their broad substrate range, i.e. the ability to use simple and complex carbohydrates including cellulose, hemicellulose and xylans (Lynd, 1989; Blumer-Schuette et al, 2008; Cordova et al, 2016); high specific substrate uptake rates and growth rates (Cordova et al, 2015; Marchant et al, 2002), which can be several-fold higher compared to mesophiles such as E. coli (Long et al, 2017a); and the ability to survive and maintain metabolic activity under harsh process conditions. Additionally, from a bioprocessing perspective, high process temperatures provide important economic benefits, including reduced cooling costs, reduced chances of contamination, easier processing of feedstocks, and the potential to integrate fermentation and product recovery (Lin and Xu, 2013; Lynd, 1989).…”