Metabolic modeling of polyhydroxybutyrate biosynthesis

Leaf, Timothy; Srienc, Friedrich

doi:10.1002/(sici)1097-0290(19980305)57:5<557::aid-bit8>3.0.co;2-f

Cited by 65 publications

(43 citation statements)

References 49 publications

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“…Above postulates are also useful tools for modelling of processes based on whey as C-source. In literature a lot of different types of mathematical models (formal kinetic, low and high structured models) are available arranged for microbia1 PHA production, but predominantly dealing with glucose, fructose, sucrose and glycerol as the C sources (Sonnleitner et al, 1979;Mulchadani et al, 1988;Raje & Srivastava, 1998;Leaf & Srienc, 1998;Katoh et al, 1999;Patwardhan & Srivastava, 2004;Franz et al, 2011). In contrast, mathematical models of processes with whey itself or with whey sugars are still rare.…”

Section: Mathematical Models For Value-added Conversion Of Wheymentioning

confidence: 99%

Whey Lactose as a Raw Material for Microbial Production of Biodegradable Polyesters

et al. 2012

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Section: Mathematical Models For Value-added Conversion Of Wheymentioning

confidence: 99%

Whey Lactose as a Raw Material for Microbial Production of Biodegradable Polyesters

et al. 2012

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“…The genome has been sequenced (http://jgi.doe .gov/JGI_microbial/html/ralstonia/ralston_homepage.html), and numerous high-cell-density processes have been reported previously (13,30). Unlike E. coli and other enterobacteria which preferentially metabolize hexose sugars through the EmbdenMeyerhof-Parnas pathway, R. eutropha preferentially uses the Entner-Douderoff pathway to metabolize hexoses (21,34). This results in a higher NADPH/NADH ratio, which causes a higher degree of biosynthetic as opposed to respiratory reducing equivalents.…”

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confidence: 99%

A Novel High-Cell-Density Protein Expression System Based on Ralstonia eutropha

Srinivasan

Barnard

Gerngross

2002

Appl Environ Microbiol

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We describe the development of a novel protein expression system based on the industrial fermentation organism Ralstonia eutropha (formerly known as Alcaligenes eutrophus) NCIMB 40124. This new system overcomes some of the shortcomings of traditional Escherichia coli-based protein expression systems, particularly the propensity of such systems to form inclusion bodies during high-level expression. Using a proteomics approach, we identified promoters that can be induced by simple process parameters or medium compositions in high-density cell culture or shake flasks, respectively. By combining newly developed molecular biological tools with a high-cell-density fermentation process, we were able to produce high levels (>1 g/liter) of soluble, active organophosphohydrolase, a model enzyme prone to inclusion body formation in E. coli.Advances in protein engineering, the completion of numerous bacterial and fungal genome sequencing projects, and the isolation of new genes from extremophiles have led to an increased number of useful proteins. However, to enable recombinant proteins to play a role in applications where larger quantities are required, such as tissue engineering or catalytic materials, production technologies that are more efficient and robust are desirable. Improving protein production is the primary goal of recombinant microbial process development and is a focus of our laboratory. Overall protein productivity can be improved by increasing the product of two variables: (i) the amount of recombinant protein per cell (specific productivity) and (ii) the amount of cell mass per unit of volume and time (cell productivity). In order to improve volumetric productivity in a cost-effective manner, recombinant proteins are often produced in high-cell-density fermentations. High-cell-density fermentations offer many advantages over traditional fermentations in that final product concentrations are higher and downtime and water usage are reduced, yet overall productivity is improved, resulting in lower setup and operating costs (3, 6).In the absence of specific folding or posttranslational modification requirements, Escherichia coli is usually the expression host of choice. E. coli-based fermentation systems produce good yields at laboratory scale; however, scale up to industrial scale, where oxygen enrichment, dialysis, and sophisticated feeding algorithms become impractical and cost prohibitive, has been challenging (22). Unlike laboratory-scale fermentors, where the above-mentioned techniques are feasible, largescale industrial fermentors are limited by mixing constraints and their ability to transfer oxygen and heat. It is well documented that oxygen-limiting conditions in E. coli result in the production of reduced carbon metabolites such as acetate, lactate, and formate (1, 20), which accumulate in high-celldensity cultures and ultimately inhibit further microbial growth. Titers of recombinant proteins in E. coli are limited by several factors including the final cell density (22), the tendency for inclusion...

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“…The PHA depolymerase enzyme is a constitutive enzyme, which means that even during PHA accumulation PHA, depolymerase is active , such that PHA polymerisation and degradation can occur simultaneously inside the cell (Uchino et al 2007 (Leaf and Srienc 1998), which also regulate β-oxidation. In this sense, simultaneous PHA synthesis and degradation are believed to give the cell an advantage under shifting environmental conditions of carbon and nutrient availability (Ren et al 2009).…”

Section: Regulation Of Pha Metabolism 221mentioning

confidence: 99%

“…In the case of PHA storage, intracellular metabolite ratios (AcCoA/CoASH and NADPH/NADP + ) have been shown to play a key regulatory role (Leaf and Srienc 1998). For instance, PHB production requires acetyl-CoA and reducing power (NADPH), whereas active biomass synthesis requires acetylCoA and ATP (and some NADPH) (Leaf and Srienc 1998).…”

Section: Considering That a Maximal Theoretical Yield Of Phb (Y Phb/smentioning

confidence: 99%

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Composition of mixed culture PHA biopolymers and implications for downstream processing

Montano-Herrera¹

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Polyhydroxyalkanoates (PHAs) are biopolymers synthesised and accumulated by most bacteria for carbon and energy storage. They have properties and applications comparable to petrochemical thermoplastics. Although PHAs are produced at high yields using pure biological cultures, the use of mixed cultures can significantly reduce the production costs and make use of waste streams to produce environmentally sustainable materials. In order to produce mixed culture PHAs of relevance for broader industry applications it is necessary to develop the ability to tailor polymers with diversified mechanical properties through understanding and controlling the monomer composition and compositional distribution.Diverse and complex PHA polymer structures have been achieved in mixed microbial cultures using time-based feeding strategies. However, PHA monomer composition and compositional distribution in PHA random and blocky copolymers are sensitive to substrate feeding history, making more complex the prediction of final PHA content and composition.In this sense, a better understanding of the changing cell physiologies that develop in response to different feeding strategies and substrates is necessary to design optimised feeding strategies and process control algorithms for PHA production.This thesis presents for the first time a comprehensive flux characterisation of monomer development during mixed culture PHA accumulation concurrent with biomass growth using Metabolic Flux Analysis (MFA). A dynamic trend in active biomass growth and in polymer composition was observed and was consistent over replicate accumulations. Monomer (3-hydroxybutyrate and 3-hydroxyvalerate) incorporation into poly(3-hydroxybutyate-co-3-hydroxyvalerate) copolymers in a pilot scale production system was evaluated based on published models that describe polymer production during PHA accumulation. However, it was found that these existing models could not describe the fluctuations in the proportion of Additionally, thermal degradation of mixed culture PHAs during melt processing was assessed by Near-Infrared (NIR) spectroscopy coupled to Multivariate Data Analysis. It was shown that, with correct pretreatment, a copolymeric product that was much more stable to extrusion processing than commercially available PHA was produced.Overall, this thesis explores both the fundamental and applied aspects of PHA production by mixed microbial cultures with concurrent active biomass growth. The use of computational iii tools to explore and help understand the underlying metabolic processes was explored.Polymer composition seems to follow a very complex regulation processes which can be described through the incorporation of more detailed reactions in current metabolic models.Furthermore, this thesis gives further insight into the fundamental properties of the materials produced and an assessment of their potential for degradation during processing.iv

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Metabolic modeling of polyhydroxybutyrate biosynthesis

Cited by 65 publications

References 49 publications

Whey Lactose as a Raw Material for Microbial Production of Biodegradable Polyesters

Whey Lactose as a Raw Material for Microbial Production of Biodegradable Polyesters

A Novel High-Cell-Density Protein Expression System Based on Ralstonia eutropha

Composition of mixed culture PHA biopolymers and implications for downstream processing

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