Simultaneous aerobic nitrogen and phosphate removal capability of novel salt-tolerant strain, Pseudomonas mendocina A4: Characterization, mechanism and application potential.

A salt-tolerant strain, Pseudomonas mendocina A4, was isolated from brackish-water ponds showing simultaneous heterotrophic nitrification-aerobic denitrification and phosphorus removal capability. The optimal conditions for nitrogen and phosphate removal of strain A4 were pH 7-8, carbon/nitrogen ratio 10, phosphorus/nitrogen ratio 0.2, temperature 30 °C, and salinity range of 0-5 % using sodium succinate as the carbon source. The nitrogen and phosphate removal efficiencies were 96-100 % and 88-96 % within 24 h, respectively. The nitrogen and phosphate removal processes were matched with the modified Gompertz model, and the underlying mechanisms were confirmed by the activities of key metabolic enzymes. Under 10 % salinity, the immobilization technology was employed to enhance the nitrogen and phosphate removal efficiencies of strain A4, achieving 87 % and 76 %, respectively. These findings highlight the potential application of strain A4 in both freshwater and marine culture wastewater treatment.

Rapid simultaneous removal of nitrogen and phosphorous by a novel isolated Pseudomonas mendocina SCZ-2.

Nitrogen (N) and phosphorous (P) removal by a single bacterium could improve the biological reaction efficiency and reduce the operating cost and complexity in wastewater treatment plants (WWTPs). Here, an isolated strain was identified as Pseudomonas mendocina SCZ-2 and showed high performance of heterotrophic nitrification (HN) and aerobic denitrification (AD) without intermediate accumulation. During the AD process, the nitrate removal efficiency and rate reached a maximum of 100% and 47.70 mg/L/h, respectively, under optimal conditions of sodium citrate as carbon source, a carbon-to-nitrogen ratio of 10, a temperature of 35 °C, and shaking a speed of 200 rpm. Most importantly, the strain SCZ-2 could rapidly and simultaneously eliminate N and P with maximum NH4+-N, NO3--N, NO2--N, and PO43--P removal rates of 14.38, 17.77, 20.13 mg N/L/h, and 2.93 mg P/L/h, respectively. Both the N and P degradation curves matched well with the modified Gompertz model. Moreover, the amplification results of functional genes, whole genome sequencing, and enzyme activity tests provided theoretical support for simultaneous N and P removal pathways. This study deepens our understanding of the role of HN-AD bacteria and provides more options for simultaneous N and P removal from actual sewage.

Efficient aerobic denitrification without nitrite accumulation by Pseudomonas mendocina HITSZ-D1 isolated from sewage sludge.

A highly efficient aerobic denitrifying microbe was isolated from sewage sludge by using a denitrifier enrichment strategy based on decreasing carbon content. The microbe was identified as Pseudomonas mendocina HITSZ-D1 (hereafter, D1). Investigation of the conditions under which D1 grew and denitrified revealed that it performed good growth and nitrate removal performance under a wide range of conditions. In particular, D1 rapidly removed all types of inorganic nitrogen without accumulation of the intermediate products nitrite and nitrous oxide. Overall, D1 showed a total nitrogen removal efficiency >96% at a C/N ratio of 8. The biotransformation modes and fates of three typical types of inorganic nitrogen were also assessed. Moreover, D1 had significantly higher denitrification efficiency and enzyme activities than other aerobic denitrifying microbes (Paracoccus denitrificans, Pseudomonas aeruginosa, and Pseudomonas putida). These results suggest that D1 has great potential for treating wastewater containing high concentrations of nitrogen.

Effects of bamboo-charcoal modified by bimetallic Fe/Pd nanoparticles on n-hexane biodegradation by bacteria Pseudomonas mendocina NX-1.

The high hydrophobicity of n-hexane is the main reason why it is difficult to be removed biologically. In this study, the effects of bamboo-charcoal modified by bimetallic Fe/Pd (BBC) on n-hexane biodegradation by Pseudomonas mendocina NX-1 (PM) was investigated. The n-hexane removal efficiency was increased in the presence of BC. The highest n-hexane removal efficiency at 90.0% was achieved at 0.05 g L-1 BCE and 3 g L-1 NH4+ under pH 7.7 and 35 °C. Additionally, protein content (45.9 µg mL-1) and negative cell surface zeta potential (-26.4 mV) were increased during biodegradation process, with PM-BBC being 43.1 µg mL-1 and 19.1 mV. Bacterial growth was improved and maximum cell surface hydrophobicity was obtained after 20 h, which was 59.4% higher than the control with PM-BBC (37.7%) or PM (16.1%), showing biodegradation products of 1-butanol and acetic acid. The results indicate that BBC improved n-hexane biodegradation efficiency by promoting bacterial growth, reducing cell zeta potential, exposing hydrophobic proteins, and increasing cell surface hydrophobicity of bacterial strain NX-1. This investigation suggests that BBC-enhanced biodegradation can be promising to treat n-hexane-containing gas.

Class II LitR serves as an effector of "short" LOV-type blue-light photoreceptor in Pseudomonas mendocina.

PmlR2, a class II LitR/CarH family transcriptional regulator, and PmSB-LOV, a "short" LOV-type blue light photoreceptor, are adjacently encoded in Pseudomonas mendocina NBRC 14162. An effector protein for the "short" LOV-type photoreceptor in Pseudomonas has not yet been identified. Here, we show that PmlR2 is an effector protein of PmSB-LOV. Transcriptional analyses revealed that the expression of genes located near pmlR2 and its homolog gene, pmlR1, was induced in response to illumination. In vitro DNA-protein binding analyses showed that recombinant PmlR2 directly binds to the promoter region of light-inducible genes. Furthermore PmSB-LOV exhibited a typical LOV-type light-induced spectral change. Gel-filtration chromatography demonstrated that the illuminated PmSB-LOV was directly associated with PmlR2, whereas non-illuminated proteins did not interact. The inhibition of PmlR2 function following PmSB-LOV binding was verified by surface plasmon resonance: the DNA-binding ability of PmlR2 was specifically inhibited in the presence of blue light-illuminated-PmSB-LOV. An In vitro transcription assay showed a dose-dependent reduction in PmlR2 repressor activity in the presence of illuminated PmSB-LOV. Overall, evidence suggests that the DNA-binding activity of PmlR2 is inhibited by its direct association with blue light-activated PmSB-LOV, enabling transcription of light-inducible promoters by RNA polymerase.

Filling in the Gaps in Metformin Biodegradation: a New Enzyme and a Metabolic Pathway for Guanylurea.

The widely prescribed pharmaceutical metformin and its main metabolite, guanylurea, are currently two of the most common contaminants in surface and wastewater. Guanylurea often accumulates and is poorly, if at all, biodegraded in wastewater treatment plants. This study describes Pseudomonas mendocina strain GU, isolated from a municipal wastewater treatment plant, using guanylurea as its sole nitrogen source. The genome was sequenced with 36-fold coverage and mined to identify guanylurea degradation genes. The gene encoding the enzyme initiating guanylurea metabolism was expressed, and the enzyme was purified and characterized. Guanylurea hydrolase, a newly described enzyme, was shown to transform guanylurea to one equivalent (each) of ammonia and guanidine. Guanidine also supports growth as a sole nitrogen source. Cell yields from growth on limiting concentrations of guanylurea revealed that metabolism releases all four nitrogen atoms. Genes encoding complete metabolic transformation were identified bioinformatically, defining the pathway as follows: guanylurea to guanidine to carboxyguanidine to allophanate to ammonia and carbon dioxide. The first enzyme, guanylurea hydrolase, is a member of the isochorismatase-like hydrolase protein family, which includes biuret hydrolase and triuret hydrolase. Although homologs, the three enzymes show distinct substrate specificities. Pairwise sequence comparisons and the use of sequence similarity networks allowed fine structure discrimination between the three homologous enzymes and provided insights into the evolutionary origins of guanylurea hydrolase.IMPORTANCE Metformin is a pharmaceutical most prescribed for type 2 diabetes and is now being examined for potential benefits to COVID-19 patients. People taking the drug pass it largely unchanged, and it subsequently enters wastewater treatment plants. Metformin has been known to be metabolized to guanylurea. The levels of guanylurea often exceed that of metformin, leading to the former being considered a "dead-end" metabolite. Metformin and guanylurea are water pollutants of emerging concern, as they persist to reach nontarget aquatic life and humans, the latter if it remains in treated water. The present study has identified a Pseudomonas mendocina strain that completely degrades guanylurea. The genome was sequenced, and the genes involved in guanylurea metabolism were identified in three widely separated genomic regions. This knowledge advances the idea that guanylurea is not a dead-end product and will allow for bioinformatic identification of the relevant genes in wastewater treatment plant microbiomes and other environments subjected to metagenomic sequencing.

Differential expression of proteins in Pseudomonas mendocina SMSKVR-3 under arsenate stress.

This study focuses on analyzing the protein expression pattern of intracellular proteins when Pseudomonas mendocina SMSKVR-3 exposed to 300 mM of arsenate to find out the proteins that are overexpressed or exclusively expressed in response to arsenate. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of protein expression at different time intervals showed the highest number of protein bands (14) that are overexpressed at 8 h of the time interval. It was also observed that treatment with at least 200 mM of As(V) is required to induce a difference in protein expression. Two-dimensional (2D)-PAGE analysis of 8-h sample exhibited 146 unique spots, 45 underexpressed, and 46 overexpressed spots in arsenate-treated sample. Based on the highest percent volume and fold change, three unique spots and one overexpressed spot were selected and analyzed by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF/TOF) mass spectrometry (MS) analysis followed by the MASCOT search. These proteins were identified as ribosome-recycling factor (20.13 kDa), polyphosphate:ADP/GDP phosphotransferase (40.88 kDa), ribonuclease P protein component (14.96 kDa) and cobalt-precorrin-5B C(1)-methyltransferase (38.43 kDa) with MASCOT score of 54, 81, 94, and 100, respectively. All of these proteins help the bacteria to overcome arsenate stress.

Bacteria involved in biodegradation of creosote PAH - A case study of long-term contaminated industrial area.

Polycyclic aromatic hydrocarbons (PAH) contained in creosote oil are particularly difficult to remove from the soil environment. Their hydrophobic character and low bioavailability to soil microorganisms affects their rate of biodegradation. This study was performed on samples of soil that were (for over forty years) subjected to contamination with creosote oil, and their metagenome and physicochemical properties were characterized. Moreover, the study was undertaken to evaluate the biodegradation of PAHs by autochthonous consortia as well as by selected bacteria strains isolated from long-term contaminated industrial soil. From among the isolated microorganisms, the most effective in biodegrading the contaminants were the strains Pseudomonas mendocina and Brevundimonas olei. They were able to degrade more than 60% of the total content of PAHs during a 28-day test. The biodegradation of these compounds using AT7 dispersant was enhanced only by Serratia marcescens strain. Moreover, the addition of AT7 improved the effectiveness of fluorene and acenaphthene biodegradation by Serratia marcescens 6-fold. Our results indicated that long-term contact with aromatic compounds induced the bacterial strains to use the PAHs as a source of carbon and energy. We observed that supplementation with surfactants does not increase the efficiency of hydrocarbon biodegradation.

Removal of nitrogen by heterotrophic nitrification-aerobic denitrification of a novel halotolerant bacterium Pseudomonas mendocina TJPU04.

Excess inorganic nitrogen in water poses a severe threat to enviroment. Removal of inorganic nitrogen by heterotrophic nitrifying-aerobic denitrifying microorganism is supposed to be a promising and applicable technology only if the removal rate can be maintained sufficiently high in real wastewater under various conditions, such as high concentration of salt and wide range of different nitrogen concentrations. Here, a new heterotrophic nitrifying-aerobic denitrifying bacterium was isolated and named as Pseudomonas mendocina TJPU04, which removes NH4+-N, NO3--N and NO2--N with average rate of 4.69, 5.60, 4.99 mg/L/h, respectively. It also maintains high nitrogen removal efficiency over a wide range of nitrogen concentrations. When concentration of NH4+-N, NO3--N and NO2--N was up to 150, 150 and 50 mg/L, 98%, 93%, and 100% removal efficiency could be obtained, respectively, after 30-h incubation under sterile condition. When it was applied under non-sterile condition, the ammonia removal efficiency was slightly lower than that under sterile condition. However, the nitrate and nitrite removal efficiencies under non-sterile condition were significantly higher than those under sterile condition. Strain TJPU04 also showed efficient nitrogen removal performance in the presence of high concentration of salt and nitrogen. In addition, the removal efficiencies of NH4+-N, NO3--N and TN in real wastewater were 91%, 52%, and 75%, respectively. These results suggest that strain TJPU04 is a promising candidate for efficient removal of inorganic nitrogen in wastewater treatment.

Screening of endogenous strong promoters for enhanced production of medium-chain-length polyhydroxyalkanoates in Pseudomonas mendocina NK-01.

Polyhydroxyalkanoate (PHA) can be produced by microorganisms from renewable resources and is regarded as a promising bioplastic to replace petroleum-based plastics. Pseudomonas mendocina NK-01 is a medium-chain-length PHA (mcl-PHA)-producing strain and its whole-genome sequence is currently available. The yield of mcl-PHA in P. mendocina NK-01 is expected to be improved by applying a promoter engineering strategy. However, a limited number of well-characterized promoters has greatly restricted the application of promoter engineering for increasing the yield of mcl-PHA in P. mendocina NK-01. In this work, 10 endogenous promoters from P. mendocina NK-01 were identified based on RNA-seq and promoter prediction results. Subsequently, 10 putative promoters were characterized for their strength through the expression of a reporter gene gfp. As a result, five strong promoters designated as P4, P6, P9, P16 and P25 were identified based on transcriptional level and GFP fluorescence intensity measurements. To evaluate whether the screened promoters can be used to enhance transcription of PHA synthase gene (phaC), the three promoters P4, P6 and P16 were separately integrated into upstream of the phaC operon in the genome of P. mendocina NK-01, resulting in the recombinant strains NKU-4C1, NKU-6C1 and NKU-16C1. As expected, the transcriptional levels of phaC1 and phaC2 in the recombinant strains were increased as shown by real-time quantitative RT-PCR. The phaZ gene encoding PHA depolymerase was further deleted to construct the recombinant strains NKU-∆phaZ-4C1, NKU-∆phaZ-6C1 and NKU-∆phaZ-16C1. The results from shake-flask fermentation indicated that the mcl-PHA titer of recombinant strain NKU-∆phaZ-16C1 was increased from 17 to 23 wt% compared with strain NKU-∆phaZ. This work provides a feasible method to discover strong promoters in P. mendocina NK-01 and highlights the potential of the screened endogenous strong promoters for metabolic engineering of P. mendocina NK-01 to increase the yield of mcl-PHA.

Morphology engineering for enhanced production of medium-chain-length polyhydroxyalkanoates in Pseudomonas mendocina NK-01.

Polyhydroxyalkanoates (PHAs) can be produced by microorganisms from renewable resources and are regarded as promising bioplastics to replace petroleum-based plastics. A medium-chain-length PHAs (mcl-PHA)-producing strain Pseudomonas mendocina NK-01 was isolated previously by our lab and its whole-genome sequence is currently available. Morphology engineering of manipulating cell morphology-related genes has been applied for enhanced accumulation of the intracellular biopolymer short-chain-length PHAs (scl-PHA). However, it has not yet been reported to improve the yield of mcl-PHA by morphology engineering so far. In this work, several well-characterized cell morphology-related genes, including the cell fission ring (Z-ring) location genes minCD, peptidoglycan degradation gene nlpD, actin-like cytoskeleton protein gene mreB, Z-ring formation gene ftsZ, and FtsZ inhibitor gene sulA, were intensively investigated for their impacts on the cell morphology and mcl-PHA accumulation by gene knockout and overexpression in P. mendocina NKU, a upp knockout mutant of P. mendocina NK-01. For a minCD knockout mutant P. mendocina NKU-∆minCD, the average cell length was obviously increased and the mcl-PHA production was improved. However, the nlpD knockout mutant had a shorter cell length and lower mcl-PHA yield compared with P. mendocina NKU. Overexpression of mreB in P. mendocina NKU resulted in spherical cells. When ftsZ was overexpressed in P. mendocina NKU, the cell division was accelerated and the mcl-PHA titer was improved. Furthermore, mreB, ftsZ, or sulA was overexpressed in P. mendocina NKU-∆minCD. Consequently, the mcl-PHA titers were all increased compared with P. mendocina NKU-∆minCD carrying the empty vector. The multiple fission pattern was finally achieved in ftsZ-overexpressing NKU-∆minCD. In this work, improved production of mcl-PHA in P. mendocina NK-01 has been achieved by morphology engineering. This work provides an alternative strategy to enhance mcl-PHA accumulation in mcl-PHA-producing strains.

Biosynthesis and characterization of a novel, biocompatible medium chain length polyhydroxyalkanoate by Pseudomonas mendocina CH50 using coconut oil as the carbon source.

This study validated the utilization of triacylglycerides (TAGs) by Pseudomonas mendocina CH50, a wild type strain, resulting in the production of novel mcl-PHAs with unique physical properties. A PHA yield of 58% dcw was obtained using 20 g/L of coconut oil. Chemical and structural characterisation confirmed that the mcl-PHA produced was a terpolymer comprising of three different repeating monomer units, 3-hydroxyoctanoate, 3-hydroxydecanoate and 3-hydroxydodecanoate or P(3HO-3HD-3HDD). Bearing in mind the potential of P(3HO-3HD-3HDD) in biomedical research, especially in neural tissue engineering, in vitro biocompatibility studies were carried out using NG108-15 (neuronal) cells. Cell viability data confirmed that P(3HO-3HD-3HDD) supported the attachment and proliferation of NG108-15 and was therefore confirmed to be biocompatible in nature and suitable for neural regeneration.

Production and recovery of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) from biodiesel liquid waste (BLW).

Polyhydroxyalkanoates (PHAs) has been paid great attention because of its useful thermoplastic properties and complete degradation in various natural environments. But, at industrial level, the successful commercialization of PHAs is limited by the high production cost due to the expensive carbon source and recovery processes. Pseudomonas mendocina PSU cultured for 72 h in mineral salts medium (MSM) containing 2% (v/v) biodiesel liquid waste (BLW) produced 79.7 wt% poly(3-hydroxybutyrate) (PHB) at 72 h. In addition, this strain produced 43.6 wt% poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) with 8.6 HV mol% at 60 h when added with 0.3% sodium propionate. The synthesized intracellular PHA granules were recovered and purified by the recently reported biological method using mealworms. The weight average molecular weight (Mw ) and number average molecular weight (Mn ) of the biologically extracted PHA were higher than that from the chloroform extraction with comparable melting temperature (Tm ) and high purity. This study has successfully established a low-cost process to synthesize PHAs from BLW and subsequently confirmed the ability of mealworms to extract PHAs from various kinds of bacterial cells.

Cyanotrophic and arsenic oxidizing activities of Pseudomonas mendocina P6115 isolated from mine tailings containing high cyanide concentration.

Mine tailings and wastewater generate man-made environments with several selective pressures, including the presence of heavy metals, arsenic and high cyanide concentrations, but severe nutritional limitations. Some oligotrophic and pioneer bacteria can colonise and grow in mine wastes containing a low concentration of organic matter and combined nitrogen sources. In this study, Pseudomonas mendocina P6115 was isolated from mine tailings in Durango, Mexico, and identified through a phylogenetic approach of 16S rRNA, gyrB, rpoB, and rpoD genes. Cell growth, cyanide consumption, and ammonia production kinetics in a medium with cyanide as sole nitrogen source showed that at the beginning, the strain grew assimilating cyanide, when cyanide was removed, ammonium was produced and accumulated in the culture medium. However, no clear stoichiometric relationship between both nitrogen sources was observed. Also, cyanide complexes were assimilated as nitrogen sources. Other phenotypic tasks that contribute to the strain's adaptation to a mine tailing environment included siderophores production in media with moderate amounts of heavy metals, arsenite and arsenate tolerance, and the capacity of oxidizing arsenite. P. mendocina P6115 harbours cioA/cioB and aoxB genes encoding for a cyanide-insensitive oxidase and an arsenite oxidase, respectively. This is the first report where P. mendocina is described as a cyanotrophic and arsenic oxidizing species. Genotypic and phenotypic tasks of P. mendocina P6115 autochthonous from mine wastes are potentially relevant for biological treatment of residues contaminated with cyanide and arsenic.

Aerobic Biodegradation of OCDD by P. Mendocina NSYSU: Effectiveness and Gene Inducement Studies.

The goals of this study were to assess the effectiveness of (1) enhancing octachlorinated dibenzo-p-dioxin (OCDD) biodegradation under aerobic conditions by Pseudomonas mendocina NSYSU (P. Mendocina NSYSU) with the addition of lecithin, and (2) inducing OCDD ring-cleavage genes by pentachlorophenol (PCP) and OCDD addition. P. Mendocina NSYSU could biodegrade OCDD via aerobic cometabolism and lecithin was used as a primary substrate. Approximately 74 and 67% of OCDD biodegradation was observed after 60 days of incubation with lecithin and glucose supplement, respectively. Lecithin was also used as the solubilization additive resulting in OCDD solubilization and enhanced bioavailability of OCDD to P. Mendocina NSYSU. Two intradiol and extradiol ring-cleavage dioxygenase genes (Pmen_0474 and Pmen_2526) were identified from gene analyses. Gene concentration was significantly enhanced after the inducement by PCP and OCDD. Higher gene inducement efficiency was obtained using PCP as the inducer, and Pmen_2526 played a more important role in OCDD biodegradation.

Production of a novel medium chain length poly(3-hydroxyalkanoate) using unprocessed biodiesel waste and its evaluation as a tissue engineering scaffold.

This study demonstrated the utilization of unprocessed biodiesel waste as a carbon feedstock for Pseudomonas mendocinaCH50, for the production of PHAs. A PHA yield of 39.5% CDM was obtained using 5% (v/v) biodiesel waste substrate. Chemical analysis confirmed that the polymer produced was poly(3-hydroxyhexanoate-co-3-hydroxyoctanoate-co-3-hydroxydecanoate-co-3-hydroxydodecanoate) or P(3HHx-3HO-3HD-3HDD). P(3HHx-3HO-3HD-3HDD) was further characterized and evaluated for its use as a tissue engineering scaffold (TES). This study demonstrated that P(3HHx-3HO-3HD-3HDD) was biocompatible with the C2C12 (myoblast) cell line. In fact, the % cell proliferation of C2C12 on the P(3HHx-3HO-3HD-3HDD) scaffold was 72% higher than the standard tissue culture plastic confirming that this novel PHA was indeed a promising new material for soft tissue engineering.

Comparative investigation on a hexane-degrading strain with different cell surface hydrophobicities mediated by starch and chitosan.

Bioremediation usually exhibits low removal efficiency toward hexane because of poor water solubility, which limits the mass transfer rate between the substrate and microorganism. This work aimed to enhance the hexane degradation rate by increasing cell surface hydrophobicity (CSH) of the degrader, Pseudomonas mendocina NX-1. The CSH of P. mendocina NX-1 was manipulated by treatment with starch and chitosan solution of varied concentrations, reaching a maximum hydrophobicity of 52%. The biodegradation of hexane conformed to the Haldane inhibition model, and the maximum degradation rate (ν max) of the cells with 52% CSH was 0.72 mg (mg cell)-1·h-1 in comparison with 0.47 mg (mg cell)-1·h-1 for cells with 15% CSH. The production of CO2 by high CSH cells was threefold higher than that by cells at 15% CSH within 30 h, and the cumulative rates of O2 consumption were 0.16 and 0.05 mL/h, respectively. High CSH was related to low negative charge carried by the cell surface and probably reduced the repulsive electrostatic interactions between hexane and microorganisms. The FT-IR spectra of cell envelopes demonstrated that the methyl chain was inversely proportional to increasing CSH values, but proteins exhibited a positive effect to CSH enhancement. The ratio of extracellular proteins and polysaccharides increased from 0.87 to 3.78 when the cells were treated with starch and chitosan, indicating their possible roles in increased CSH.

Biosynthesis, biotechnological production, and applications of glucosylglycerols.

Glucosylglycerols (GGs) are known as compatible solutes accumulated by some bacteria including cyanobacteria as well as higher plants for their adaptations to salt or desiccation stresses. Since being identified in Japanese sake, their physiological effects and potential applications on human health cares have been explored in the following 15 years. Several different synthesis methods have been successively developed for the production of GGs. However, the efficiency of GG synthesis, especially biological synthesis, is still low. With the recent advances in genome sequencing and synthetic biology tools, systematical screening of enzyme candidates and metabolic engineering approaches is necessary for improving GG synthesis efficiency. In this review, we will summarize GG structure information, protective effects on human skin and digestive system as well as industrial enzymes, together with their synthesis by chemical, enzymatic, and biological in vivo approaches in detail, and provide some prospects on improving GG production.

Efficient production of polyhydroxyalkanoates (PHAs) from Pseudomonas mendocina PSU using a biodiesel liquid waste (BLW) as the sole carbon source.

Conditions for the optimal production of polyhydroxyalkanoate (PHA) by Pseudomonas mendocina PSU using a biodiesel liquid waste (BLW) were determined by response surface methodology. These were an initial carbon to nitrogen ratio (C/N) of 40 (mole/mole), an initial pH of 7.0, and a temperature of 35 °C. A biomass and PHA concentration of 3.65 g/L and about 2.6 g/L (77% DCW), respectively, were achieved in a growth associated process using 20 g/L glycerol in the BLW after 36 h of exponential growth. The PHA monomer compositions were 3HB (3-hydroxybutyrate), a short-chain-length-PHA, and the medium-chain-length-PHA e.g. 3-hydroxyoctanoate and 3-hydroxydecanoate. Both the phbC and phaC genes were characterized. The phbC enzyme had not been previously detected in a Pseudomonas mendocina species. A 2.15 g/L of an exopolysaccharide, alginate, was also produced with a similar composition to that of other Pseudomonas species.

Enhancement of medium-chain-length polyhydroxyalkanoates biosynthesis from glucose by metabolic engineering in Pseudomonas mendocina.

OBJECTIVES: To enhance the biosynthesis of medium-chain-length polyhydroxyalkanoates (PHAMCL) from glucose in Pseudomonas mendocina NK-01, metabolic engineering strategies were used to block or enhance related pathways. RESULTS: Pseudomonas mendocina NK-01 produces PHAMCL from glucose. Besides the alginate oligosaccharide biosynthetic pathway proved by our previous study, UDP-D-glucose and dTDP-L-rhamnose biosynthetic pathways were identified. These might compete for glucose with the PHAMCL biosynthesis. First, the alg operon, galU and rmlC gene were deleted one by one, resulting in NK-U-1(∆alg), NK-U-2 (∆alg∆galU), NK-U-3(alg∆galU∆rmlC). After fermentation for 36 h, the cell dry weight (CDW) and PHAMCL production of these strains were determined. Compared with NK-U: 1) NK-U-1 produced elevated CDW (from 3.19 ± 0.16 to 3.5 ± 0.11 g/l) and equal PHAMCL (from 0.78 ± 0.06 to 0.79 ± 0.07 g/l); 2) NK-U-2 produced more CDW (from 3.19 ± 0.16 to 3.55 ± 0.23 g/l) and PHAMCL (from 0.78 ± 0.06 to 1.05 ± 0.07 g/l); 3) CDW and PHAMCL dramatically decreased in NK-U-3 (1.53 ± 0.21 and 0.41 ± 0.09 g/l, respectively). Additionally, the phaG gene was overexpressed in strain NK-U-2. Although CDW of NK-U-2/phaG decreased to 1.29 ± 0.2 g/l, PHA titer (%CDW) significantly increased from 24.5 % up to 51.2 %. CONCLUSION: The PHAMCL biosynthetic pathway was enhanced by blocking branched metabolic pathways in combination with overexpressing phaG gene.