Modelling cell growth and polyhydroxyalkanoate (PHA) polymer synthesis by Pseudomonas putida LS46 under oxygen-limiting conditions

2021 ◽  
Vol 11 ◽  
Author(s):  
Shabnam Sharifyazd ◽  
Masoud Asadzadeh ◽  
David B. Levin
Keyword(s):  
Cell Growth ◽  
Pseudomonas Putida ◽  
Logistic Model ◽  
Absolute Error ◽  
Scale Production ◽  
Batch Mode ◽  
Medium Chain Length ◽  
Plastic Materials ◽  
Oxygen Excess ◽  
Pha Production

Background: Polyhydroxyalkanoates (PHAs) are biodegradable, biocompatible, and non-toxic polymers synthesized by bacteria that may be used to displace some petroleum-based plastic materials. One of the major barriers to the commercialization of PHA biosynthesis is the high cost of production. Objective: Oxygen-limitation is known to greatly influence bacterial cell growth and PHA production. In this study, the growth and synthesis of medium chain length PHAs (mcl-PHAs) by Pseudomonas putida LS46, cultured in batch-mode with octanoic acid, under oxygen-limited conditions, was modeled. Methods: Four models, including the Monod model, incorporated Leudeking–Piret (MLP), the Moser model incorporated Leudeking–Piret (Moser-LP), the Logistic model incorporated Leudeking–Piret (LLP), and the Modified Logistic model incorporated Leudeking–Piret (MLLP) were investigated. Kinetic parameters of each model were calibrated by using the multi-objective optimization algorithm, Pareto Archived Dynamically Dimensioned Search (PA-DDS), by minimizing the sum of absolute error (SAE) for PHA production and growth simultaneously. Results and Conclusions: Among the four models, MLP and Moser-LP models adequately represented the experimental data for oxygen-limited conditions. However, the MLP and Moser-LP models could not adequately simulate PHA production under oxygen-excess conditions. Modeling cell growth and PHA will assist in the development of a strategy for industrial-scale production.

Kinetics of medium-chain-length polyhydroxyalkanoate production by a novel isolate of Pseudomonas putida LS46

2012 ◽  
Vol 58 (8) ◽  
pp. 982-989 ◽  
Author(s):  
Parveen K. Sharma ◽  
Jilagamazhi Fu ◽  
Nazim Cicek ◽  
Richard Sparling ◽  
David B. Levin
Keyword(s):  
Pseudomonas Putida ◽  
Chain Length ◽  
Saturated Fatty Acids ◽  
Dry Mass ◽  
Nucleotide Sequence Analysis ◽  
Glucose Medium ◽  
Medium Chain ◽  
Medium Chain Length ◽  
Pha Production ◽  
Nitrogen Excess

Six bacteria that synthesize medium-chain-length polyhydroxyalkanoates (mcl-PHAs) were isolated from sewage sludge and hog barn wash and identified as strains of Pseudomonas and Comamonas by 16S rDNA gene sequencing. One isolate, Pseudomonas putida LS46, showed good PHA production (22% of cell dry mass) in glucose medium, and it was selected for further studies. While it is closely related to other P. putida strains (F1, KT2440, BIRD-1, GB-1, S16, and W619), P. putida LS46 was genetically distinct from these other strains on the basis of nucleotide sequence analysis of the cpn60 gene hypervariable region. PHA production was detected as early as 12 h in both nitrogen-limited and nitrogen-excess conditions. The increase in PHA production after 48 h was higher in nitrogen-limited cultures than in nitrogen-excess cultures. Pseudomonas putida LS46 produced mcl-PHAs when cultured with glucose, glycerol, or C6–C14 saturated fatty acids as carbon sources, and mcl-PHAs accounted for 56% of the cell dry mass when cells were batch cultured in medium containing 20 mmol/L octanoate. Although 3-hydroxydecanoate was the major mcl-PHA monomer (58.1–68.8 mol%) in P. putida LS46 cultured in glucose medium, 3-hydroxyoctanoate was the major monomer produced in octanoate medium (88 mol%).

scholarly journals Simultaneous Improvements ofPseudomonasCell Growth and Polyhydroxyalkanoate Production from a Lignin Derivative for Lignin-Consolidated Bioprocessing

2018 ◽  
Vol 84 (18) ◽  
Author(s):  
Xiaopeng Wang ◽  
Lu Lin ◽  
Junde Dong ◽  
Juan Ling ◽  
Wanpeng Wang ◽  
...  
Keyword(s):  
Cell Growth ◽  
Pseudomonas Putida ◽  
Consolidated Bioprocessing ◽  
Acyl Carrier Protein ◽  
Dry Weight ◽  
Content Type ◽  
Strong Promoter ◽  
Molecular Features ◽  
Pha Production ◽  
Key Genes

ABSTRACTCell growth and polyhydroxyalkanoate (PHA) biosynthesis are two key traits in PHA production from lignin or its derivatives. However, the links between them remain poorly understood. Here, the transcription levels of key genes involved in PHA biosynthesis were tracked inPseudomonas putidastrain A514 grown on vanillic acid as the sole carbon source under different levels of nutrient availability. First, enoyl-coenzyme A (CoA) hydratase (encoded byphaJ4) is stress induced and likely to contribute to PHA synthesis under nitrogen starvation conditions. Second, much higher expression levels of 3-hydroxyacyl-acyl carrier protein (ACP) thioesterase (encoded byphaG) and long-chain fatty acid-CoA ligase (encoded byalkK) under both high and low nitrogen (N) led to the hypothesis that they likely not only have a role in PHA biosynthesis but are also essential to cell growth. Third, 40 mg/liter PHA was synthesized by strain AphaJ4C1(overexpression ofphaJ4andphaC1in strain A514) under low-N conditions, in contrast to 23 mg/liter PHA synthesized under high-N conditions. Under high-N conditions, strain AalkKphaGC1(overexpression ofphaG,alkK, andphaC1in A514) produced 90 mg/liter PHA with a cell dry weight of 667 mg/liter, experimentally validating our hypothesis. Finally, further enhancement in cell growth (714 mg/liter) and PHA titer (246 mg/liter) was achieved in strain Axyl_alkKphaGC1via transcription level optimization, which was regulated by an inducible strong promoter with its regulator, XylR-PxylA, from the xylose catabolic gene cluster of the A514 genome. This study reveals genetic features of genes involved in PHA synthesis from a lignin derivative and provides a novel strategy for rational engineering of these two traits, laying the foundation for lignin-consolidated bioprocessing.IMPORTANCEWith the recent advances in processing carbohydrates in lignocellulosics for bioproducts, almost all biological conversion platforms result in the formation of a significant amount of lignin by-products, representing the second most abundant feedstock on earth. However, this resource is greatly underutilized due to its heterogeneity and recalcitrant chemical structure. Thus, exploiting lignin valorization routes would achieve the complete utilization of lignocellulosic biomass and improve cost-effectiveness. The culture conditions that encourage cell growth and polyhydroxyalkanoate (PHA) accumulation are different. Such an inconsistency represents a major hurdle in lignin-to-PHA bioconversion. In this study, we traced and compared transcription levels of key genes involved in PHA biosynthesis pathways inPseudomonas putidaA514 under different nitrogen concentrations to unveil the unusual features of PHA synthesis. Furthermore, an inducible strong promoter was identified. Thus, the molecular features and new genetic tools reveal a strategy to coenhance PHA production and cell growth from a lignin derivative.

Evaluation of medium-chain-length polyhydroxyalkanoate production byPseudomonas putidaLS46 using biodiesel by-product streams

2014 ◽  
Vol 60 (7) ◽  
pp. 461-468 ◽  
Author(s):  
Jilagamazhi Fu ◽  
Umesh Sharma ◽  
Richard Sparling ◽  
Nazim Cicek ◽  
David B. Levin
Keyword(s):  
Cell Growth ◽  
Chain Length ◽  
Low Cost ◽  
Nitrogen Concentration ◽  
Carbon Sources ◽  
Dry Mass ◽  
Experimental Conditions ◽  
Medium Chain ◽  
Medium Chain Length ◽  
Pha Production

Medium-chain-length polyhydroxyalkanoate (mcl-PHA) production by Pseudomonas putida LS46 was analyzed in shake-flask-based batch reactions, using pure chemical-grade glycerol (PG), biodiesel-derived “waste” glycerol (WG), and biodiesel-derived “waste” free fatty acids (WFA). Cell growth, substrate consumption, mcl-PHA accumulation within the cells, and the monomer composition of the synthesized biopolymers were monitored. The patterns of mcl-PHA synthesis in P. putida LS46 cells grown on PG and WG were similar but differed from that of cells grown with WFA. Polymer accumulation in glycerol-based cultures was stimulated by nitrogen limitation and plateaued after 48 h in both PG and WG cultures, with a total accumulation of 17.9% cell dry mass and 16.3% cell dry mass, respectively. In contrast, mcl-PHA synthesis was independent of nitrogen concentration in P. putida LS46 cells cultured with WFA, which accumulated to 29% cell dry mass. In all cases, the mcl-PHAs synthesized consisted primarily of 3-hydroxyoctanoate (C8) and 3-hydroxydecanoate (C10). WG and WFA supported similar or greater cell growth and mcl-PHA accumulation than PG under the experimental conditions used. These results suggest that biodiesel by-product streams could be used as low-cost carbon sources for sustainable mcl-PHA production.

scholarly journals Draft Genome Sequence of Medium-Chain-Length Polyhydroxyalkanoate-Producing Pseudomonas putida Strain LS46

2013 ◽  
Vol 1 (2) ◽  
Author(s):  
P. K. Sharma ◽  
J. Fu ◽  
X. Zhang ◽  
B. W. Fristensky ◽  
K. Davenport ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Chain Length ◽  
Genome Sequence ◽  
Draft Genome ◽  
Draft Genome Sequence ◽  
Medium Chain ◽  
Medium Chain Length

scholarly journals Quantitative ‘Omics Analyses of Medium Chain Length Polyhydroxyalkanaote Metabolism in Pseudomonas putida LS46 Cultured with Waste Glycerol and Waste Fatty Acids

PLoS ONE ◽  
2015 ◽  
Vol 10 (11) ◽  
pp. e0142322 ◽  
Author(s):  
Jilagamazhi Fu ◽  
Parveen Sharma ◽  
Vic Spicer ◽  
Oleg V. Krokhin ◽  
Xiangli Zhang ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Pseudomonas Putida ◽  
Chain Length ◽  
Medium Chain ◽  
Waste Glycerol ◽  
Medium Chain Length

scholarly journals Molecular Basis of Medium-Chain Length-PHA Metabolism of Pseudomonas putida

2020 ◽  
pp. 89-114
Author(s):  
Maria Tsampika Manoli ◽  
Natalia Tarazona ◽  
Aranzazu Mato ◽  
Beatriz Maestro ◽  
Jesús M. Sanz ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Chain Length ◽  
Molecular Basis ◽  
Medium Chain ◽  
Medium Chain Length

The Scalability of Wet Ball Milling for The Production of Nanosuspensions

2019 ◽  
Vol 7 (2) ◽  
pp. 147-161 ◽  
Author(s):  
Maria L.A.D. Lestari ◽  
Rainer H. Müller ◽  
Jan P. Möschwitzer
Keyword(s):  
Ball Milling ◽  
Large Scale ◽  
Milling Time ◽  
High Energy ◽  
Small Scale ◽  
Particle Sizes ◽  
Scale Production ◽  
Batch Mode ◽  
Pharmaceutical Ingredients ◽  
Large Scale Production

Background: Miniaturization of nanosuspensions preparation is a necessity in order to enable proper formulation screening before nanosizing can be performed on a large scale. Ideally, the information generated at small scale is predictive for large scale production. Objective: This study was aimed to investigate the scalability when producing nanosuspensions starting from a 10 g scale of nanosuspension using low energy wet ball milling up to production scales of 120 g nanosuspension and 2 kg nanosuspension by using a standard high energy wet ball milling operated in batch mode or recirculation mode, respectively. Methods: Two different active pharmaceutical ingredients, i.e. curcumin and hesperetin, have been used in this study. The investigated factors include the milling time, milling speed, and the type of mill. Results: Comparable particle sizes of about 151 nm to 190 nm were obtained for both active pharmaceutical ingredients at the same milling time and milling speed when the drugs were processed at 10 g using low energy wet ball milling or 120 g using high energy wet ball milling in batch mode, respectively. However, an adjustment of the milling speed was needed for the 2 kg scale produced using high energy wet ball milling in recirculation mode to obtain particle sizes comparable to the small scale process. Conclusion: These results confirm in general, the scalability of wet ball milling as well as the suitability of small scale processing in order to correctly identify the most suitable formulations for large scale production using high energy milling.

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