Model‐based high cell density cultivation of <i>Rhodospirillum rubrum</i> under respiratory dark conditions

Zeiger, Lisa; Grammel, Hartmut

doi:10.1002/bit.22589

Cited by 23 publications

(33 citation statements)

References 27 publications

(29 reference statements)

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“…We reasoned that, if product (fatty acid) formation is tied to cell number, then increasing culture density should increase the culture productivity. We opted to use a fed-batch bioreactor (31–33) to test this hypothesis, since it can bypass the negative impacts of high (toxicity) or low (limitation) nutrient availability.…”

Section: Resultsmentioning

confidence: 99%

Mutations That Alter the Bacterial Cell Envelope Increase Lipid Production

Lemmer

Zhang

Langer

et al. 2017

mBio

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Lipids from microbes offer a promising source of renewable alternatives to petroleum-derived compounds. In particular, oleaginous microbes are of interest because they accumulate a large fraction of their biomass as lipids. In this study, we analyzed genetic changes that alter lipid accumulation in Rhodobacter sphaeroides. By screening an R. sphaeroides Tn5 mutant library for insertions that increased fatty acid content, we identified 10 high-lipid (HL) mutants for further characterization. These HL mutants exhibited increased sensitivity to drugs that target the bacterial cell envelope and changes in shape, and some had the ability to secrete lipids, with two HL mutants accumulating ~60% of their total lipids extracellularly. When one of the highest-lipid-secreting strains was grown in a fed-batch bioreactor, its lipid content was comparable to that of oleaginous microbes, with the majority of the lipids secreted into the medium. Based on the properties of these HL mutants, we conclude that alterations of the cell envelope are a previously unreported approach to increase microbial lipid production. We also propose that this approach may be combined with knowledge about biosynthetic pathways, in this or other microbes, to increase production of lipids and other chemicals.

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Section: Resultsmentioning

confidence: 99%

Mutations That Alter the Bacterial Cell Envelope Increase Lipid Production

Lemmer

Zhang

Langer

et al. 2017

mBio

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“…The model provides a rationale for explaining various experimental findings and several new testable hypotheses could be formulated. Furthermore, as PNSB are promising candidates for biotechnical applications including the production of biohydrogen, biopolymers, or porphyrins [4,10,49-53], our model could become a valuable tool for the in silico design of genetically-engineered bacterial strains.…”

Section: Discussionmentioning

confidence: 99%

Metabolic network modeling of redox balancing and biohydrogen production in purple nonsulfur bacteria

2011

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BackgroundPurple nonsulfur bacteria (PNSB) are facultative photosynthetic bacteria and exhibit an extremely versatile metabolism. A central focus of research on PNSB dealt with the elucidation of mechanisms by which they manage to balance cellular redox under diverse conditions, in particular under photoheterotrophic growth.ResultsGiven the complexity of the central metabolism of PNSB, metabolic modeling becomes crucial for an integrated analysis of the accumulated biological knowledge. We reconstructed a stoichiometric model capturing the central metabolism of three important representatives of PNSB (Rhodospirillum rubrum, Rhodobacter sphaeroides and Rhodopseudomonas palustris). Using flux variability analysis, the model reveals key metabolic constraints related to redox homeostasis in these bacteria. With the help of the model we can (i) give quantitative explanations for non-intuitive, partially species-specific phenomena of photoheterotrophic growth of PNSB, (ii) reproduce various quantitative experimental data, and (iii) formulate several new hypotheses. For example, model analysis of photoheterotrophic growth reveals that - despite a large number of utilizable catabolic pathways - substrate-specific biomass and CO2 yields are fixed constraints, irrespective of the assumption of optimal growth. Furthermore, our model explains quantitatively why a CO2 fixing pathway such as the Calvin cycle is required by PNSB for many substrates (even if CO2 is released). We also analyze the role of other pathways potentially involved in redox metabolism and how they affect quantitatively the required capacity of the Calvin cycle. Our model also enables us to discriminate between different acetate assimilation pathways that were proposed recently for R. sphaeroides and R. rubrum, both lacking the isocitrate lyase. Finally, we demonstrate the value of the metabolic model also for potential biotechnological applications: we examine the theoretical capabilities of PNSB for photoheterotrophic hydrogen production and identify suitable genetic interventions to increase the hydrogen yield.ConclusionsTaken together, the metabolic model (i) explains various redox-related phenomena of the versatile metabolism of PNSB, (ii) delivers new hypotheses on the operation and relevance of several metabolic pathways, and (iii) holds significant potential as a tool for rational metabolic engineering of PNSB in biotechnological applications.

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“…However, much higher product yields (up to 4,300 mg liter Ϫ1 ) are listed in a review on biotechnological GSH production (20). In combination with recently established high-cell-density cultivation of R. rubrum (36), the data in Table 1 can be extrapolated to theoretical total yields of GSH of ϳ130 mg liter Ϫ1 intracellularly and 221 mg liter Ϫ1 excreted into the environment. Optimization of strains and medium conditions might be used to further increase the productivity.…”

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A Glutathione Redox Effect on Photosynthetic Membrane Expression in Rhodospirillum rubrum

Carius¹,

Henkel²,

Grammel³

2011

J Bacteriol

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The formation of intracytoplasmic photosynthetic membranes by facultative anoxygenic photosynthetic bacteria has become a prime example for exploring redox control of gene expression in response to oxygen and light. Although a number of redox-responsive sensor proteins and transcription factors have been characterized in several species during the last several years in some detail, the overall understanding of the metabolic events that determine the cellular redox environment and initiate redox signaling is still poor. In the present study we demonstrate that in Rhodospirillum rubrum, the amount of photosynthetic membranes can be drastically elevated by external supplementation of the growth medium with the low-molecular-weight thiol glutathione. Neither the widely used reductant dithiothreitol nor oxidized glutathione caused the same response, suggesting that the effect was specific for reduced glutathione. By determination of the extracellular and intracellular glutathione levels, we correlate the GSH/GSSG redox potential to the expression level of photosynthetic membranes. Possible regulatory interactions with periplasmic, membrane, and cytosolic proteins are discussed. Furthermore, we found that R. rubrum cultures excrete substantial amounts of glutathione to the environment.It is well established that the thiol-containing tripeptide glutathione (␥-Glu-Cys-Gly [GSH]) is the most abundant redox buffer in all eukaryotic and in many prokaryotic cells and that GSH is critically important for the defense of oxidative stress which inevitably is associated with respiratory life. Various functions and mechanisms where GSH is involved in bacteria are summarized in a recent review (22). During many of these reactions, two molecules of GSH are oxidized to glutathione disulfide (GSSG), and the reduced form has to be recycled by the enzyme glutathione reductase at the expense of NADPH. A signaling role for GSH, however, is thus far not readily established. In bacteria, the reversible formation of intramolecular or intermolecular disulfide bonds of cysteines of regulatory proteins appears to be a common motif in molecular redox switches. For example, the ArcB sensor of Escherichia coli or the PpsR-and RegB-type regulators in facultative anoxygenic photosynthetic bacteria undergo reversible thiol/disulfide switches when exposed to different redox conditions in vitro (21, 32). The latter group of bacteria (Rhodospirillaceae) has contributed significantly to the recent paradigms of redox signaling and control of gene expression due to its facultative phototrophic lifestyle, which allows these bacteria to adapt their energy metabolism to the redox conditions of the environment. Under conditions of limiting oxygen, the Rhodospirillaceae induce the biosynthesis of an extensive system of pigmented intracytoplasmic membranes (ICM) which harbor photosynthetic reaction centers and light-harvesting complexes. The major environmental factor that determines the amount of ICM is the availability of oxygen, since aerobic growth conditio...

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Model‐based high cell density cultivation of Rhodospirillum rubrum under respiratory dark conditions

Cited by 23 publications

References 27 publications

Mutations That Alter the Bacterial Cell Envelope Increase Lipid Production

Mutations That Alter the Bacterial Cell Envelope Increase Lipid Production

Metabolic network modeling of redox balancing and biohydrogen production in purple nonsulfur bacteria

A Glutathione Redox Effect on Photosynthetic Membrane Expression in Rhodospirillum rubrum

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