Glucose Assimilation Research Articles

Transcriptome and Metabolome Profiling of a Novel Isolate Chlorella sorokiniana G32 (Chlorophyta) Displaying Enhanced Starch Accumulation at High Growth Rate Under Mixotrophic Condition.

Chlorella sorokiniana is one of the most productive microalgal species with a high potential for the production of biofuels and other high value-added molecules. Many studies have focused on its capability of mixotrophic growth using reduced organic carbon and growth pattern shift between autotrophic and mixotrophic conditions. In this study, we investigated growth patterns of a novel isolate, C. sorokiniana G32, under mixotrophic growth conditions supplemented with a low level (1.25 g L–1) and a high level (5 g L–1) of glucose. Physiological, transcriptomic (i.e., RNA-seq), and metabolomic (i.e., LC-MS/MS) methods were used. We showed that peak growth based on OD680nm absorbance is ∼4-fold higher with high glucose vs. low glucose supplementation. Photosynthetic efficiency (Fv/Fm) in G32 mixotrophic cultures with high or low glucose supplementation remains identical to that of G32 phototrophic growth. We also found that the conversion rate between absorbance-based cell density and cell dry weight with high glucose supplementation was lower than with low glucose. This suggests that more cell biomass is produced under high glucose treatment than with low glucose. The result was confirmed via sucrose density gradient centrifugation. It is likely that accumulation of high concentration of starch may account for this effect. Transcriptomic analysis of G32 cultures (i.e., via RNA-seq) in response to reciprocal change of glucose levels reveals that expression of a subset of differentially expressed genes (DEGs) is correlated with the amount of glucose supplementation. These DEGs are designated as glucose-specific responsive (GSR) genes. GSR genes are enriched for a number of energy metabolic pathways. Together with metabolomics data (i.e., LC-MS/MS), we show that under high-level supplementation, glucose is preferentially oxidized through an oxidative pentose phosphate pathway. Collectively, our results indicate the mechanism of regulation of glucose assimilation and energy metabolism in G32 under mixotrophic conditions with different levels of glucose supplementation revealed by transcriptomic and metabolomic analyses. We propose that C. sorokiniana G32 has the potential for the production of high value-added molecules.

Histone H4 lysine 16 acetylation controls central carbon metabolism and diet-induced obesity in mice

Noncommunicable diseases (NCDs) account for over 70% of deaths world-wide. Previous work has linked NCDs such as type 2 diabetes (T2D) to disruption of chromatin regulators. However, the exact molecular origins of these chronic conditions remain elusive. Here, we identify the H4 lysine 16 acetyltransferase MOF as a critical regulator of central carbon metabolism. High-throughput metabolomics unveil a systemic amino acid and carbohydrate imbalance in Mof deficient mice, manifesting in T2D predisposition. Oral glucose tolerance testing (OGTT) reveals defects in glucose assimilation and insulin secretion in these animals. Furthermore, Mof deficient mice are resistant to diet-induced fat gain due to defects in glucose uptake in adipose tissue. MOF-mediated H4K16ac deposition controls expression of the master regulator of glucose metabolism, Pparg and the entire downstream transcriptional network. Glucose uptake and lipid storage can be reconstituted in MOF-depleted adipocytes in vitro by ectopic Glut4 expression, PPARγ agonist thiazolidinedione (TZD) treatment or SIRT1 inhibition. Hence, chronic imbalance in H4K16ac promotes a destabilisation of metabolism triggering the development of a metabolic disorder, and its maintenance provides an unprecedented regulatory epigenetic mechanism controlling diet-induced obesity.

Biochemical and genomic identification of novel thermophilic Bacillus licheniformis strains YNP1-TSU, YNP2-TSU, and YNP3-TSU with potential in 2,3-butanediol production from non-sterile food waste fermentation

In the U.S., 30–40% of all food produced ends up as waste. By taking advantage of biological processes this waste nutrients can be converted into value-added 2,3-butanediol (2,3-BDO). However, food waste contains live bacteria and fungi which must first be inactivated, a process typically done with autoclaving under high pressure steam. If contaminants are not removed they can out-compete added industrial microbes for nutrients, resulting in lower yields of bio-products. Unfortunately, sterilization methods such as autoclaving greatly increase production time, and increase energy consumption. This creates a demand for novel microorganisms that can out-compete food waste microbiota by natural means without the need for sterilization of food waste. For this study, several Bacillus licheniformis isolates from Yellowstone National Park were investigated for their potential use in the 2,3-BDO industry. B. licheniformis species are a strain of industrial importance as of late, due to the wide metabolic functions and sustainable growth conditions above 45 °C. Whole genomic sequencing data and biochemical classification of B. licheniformis YNP1-TSU, B. licheniformis YNP2-TSU, and B. licheniformis YNP3-TSU were assessed and revealed a wide assortment of anti-microbial peptides and antibodies as well as catabolic genes for assimilation of glucose, fructose, galactose, xylose, arabinose, mannose, starch and sucrose, and linoleic acid. All three B. licheniformis strains had an optimal growth temperature of 50 °C, and were identified as high 2,3-BDO producers. Each strain YNP1-TSU, YNP2-TSU, and YNP3-TSU yielded 0.44 g/g, 0.45 g/g, and 0.43 g/g from mixed sugars found in non-sterilized food waste, respectively.

Activity-Based Protein Profiling of Retaining α-Amylases in Complex Biological Samples.

Amylases are key enzymes in the processing of starch in many kingdoms of life. They are important catalysts in industrial biotechnology where they are applied in, among others, food processing and the production of detergents. In man amylases are the first enzymes in the digestion of starch to glucose and arguably also the preferred target in therapeutic strategies aimed at the treatment of type 2 diabetes patients through down-tuning glucose assimilation. Efficient and sensitive assays that report selectively on retaining amylase activities irrespective of the nature and complexity of the biomaterial studied are of great value both in finding new and effective human amylase inhibitors and in the discovery of new microbial amylases with potentially advantageous features for biotechnological application. Activity-based protein profiling (ABPP) of retaining glycosidases is inherently suited for the development of such an assay format. We here report on the design and synthesis of 1,6-epi-cyclophellitol-based pseudodisaccharides equipped with a suite of reporter entities and their use in ABPP of retaining amylases from human saliva, murine tissue as well as secretomes from fungi grown on starch. The activity and efficiency of the inhibitors and probes are substantiated by extensive biochemical analysis, and the selectivity for amylases over related retaining endoglycosidases is validated by structural studies.

De novo transcriptome analysis of Chlorella sorokiniana: effect of glucose assimilation, and moderate light intensity

Chlorella can produce an unusually wide range of metabolites under various nutrient availability, carbon source, and light availability. Glucose, an essential molecule for the growth of microorganisms, also contributes significantly to the metabolism of various metabolic compounds produced by Chlorella. In addition, manipulation of light intensity also induces the formation of secondary metabolites such as pigments, and carotenoids in Chlorella. This study will focus on the effect of glucose addition, and moderate light on the regulation of carotenoid, lipid, starch, and other key metabolic pathways in Chlorella sorokiniana. To gain knowledge about this, we performed transcriptome profiling on C. sorokiniana strain NIES-2168 in response to moderate light stress supplemented with glucose under mixotrophic conditions. A total of 60,982,352 raw paired-end (PE) reads 100 bp in length was obtained from both normal, and mixotrophic samples of C. sorokiniana. After pre-processing, 93.63% high-quality PE reads were obtained, and 18,310 predicted full-length transcripts were assembled. Differential gene expression showed that a total of 937, and 1124 genes were upregulated, and downregulated in mixotrophic samples, respectively. Transcriptome analysis revealed that the mixotrophic condition caused upregulation of genes involved in carotenoids production (specifically lutein biosynthesis), fatty acid biosynthesis, TAG accumulation, and the majority of the carbon fixation pathways. Conversely, starch biosynthesis, sucrose biosynthesis, and isoprenoid biosynthesis were downregulated. Novel insights into the pathways that link the enhanced production of valuable metabolites (such as carotenoids in C. sorokiniana) grown under mixotrophic conditions is presented.

Inhibition of glucose assimilation in Auxenochlorella protothecoides by light

BackgroundThe yield of microalgae biomass is the key to affect the accumulation of fatty acids. A few microalgae can assimilate organic carbon to improve biomass yield. In mixotrophic cultivation, microalgae can use organic carbon source and light energy simultaneously. The preference of the main energy source by microalgae determines the biomass yield. Auxenochlorella protothecoides is an oleaginous mixotrophic microalga that can efficiently assimilate glucose and accumulate a large amount of biomass and fatty acids. The current study focused on the effect of light on the growth and glucose assimilation of A. protothecoides.ResultsIn this study, we found that the uptake and metabolism of glucose in A. protothecoides could be inhibited by light, resulting in a reduction of biomass growth and lipid accumulation. We employed comparative proteomics to study the influence of light on the regulation of glucose assimilation in A. protothecoides. Proteomics revealed that proteins involving in gene translation and photosynthesis system were up-regulated in the light, such as ribulose-phosphate 3-epimerase and phosphoribulokinase. Calvin cycle-related proteins were also up-regulated, suggesting that light may inhibit glucose metabolism by enhancing the production of glyceraldehyde-3-phosphate (G3P) in the Calvin cycle. In addition, the redox homeostasis-related proteins such as thioredoxin reductase were up-regulated in the light, indicating that light may regulate glucose uptake by changing the redox balance. Moreover, the increase of NADH levels and redox potential of the medium under illumination might inhibit the activity of the glucose transport system and subsequently reduce glucose uptake.ConclusionsA theoretical model of how glucose assimilation in A. protothecoides is negatively influenced by light was proposed, which will facilitate further studies on the complex mechanisms underlying the transition from autotrophy to heterotrophy for improving biomass accumulation.

Robust production of pigment-free pullulan from lignocellulosic hydrolysate by a new fungus co-utilizing glucose and xylose

Cost-efficient production of pullulan is of great importance but remains challenging due largely to the high-cost carbon sources. Lignocellulosic biomass is considered an alternative carbon source for industrial pullulan production, while new fungus producers that co-utilizing lignocellulose-derived glucose and xylose are required. In this study, a new fungus Aureobasidium melanogenum TN2-1-2 showed simultaneously assimilation of glucose and xylose and could produce pigment-free pullulan due to its deficiency in melanin synthesis. The ability of TN2-1-2 producing pullulan was remarkably robust in the presence of varying glucose to xylose ratios and ionic salt concentrations. Furthermore, condensed lignocellulosic hydrolysates obtained by consolidated biosaccharification was used as the pullulan production medium without supplying any nutrients, and pigment-free pullulan was produced by TN2-1-2 with the titer and yield of 55.1 g/L and 0.5 gPullulan/gCBS hydrolysate, respectively. Hence, this work provides a potential industrial pullulan producer TN2-1-2 and new insight into the lignocellulose bioconversion to pullulan.

Prevotella histicola treatment reduces arthritic pain and partially normalizes gut microbiota and metabolites

Abstract We have previously shown that treatment with a novel human commensal, Prevotella histicola, isolated from a duodenal biopsy, can protect HLA-DQ8 mice from collagen induced arthritis (CIA), a model for RA, by modulating the systemic immune response. The present study analyzed the functional and mechanistic impact of P. histicola by comparing pain, motor function, gut microbiota and metabolites in treated and non-treated arthritic DQ8 mice. P. histicola treatment reduced the arthritis-associated pain by improving the motor function when compared to control mice. Analysis of synovium showed that P. histicola treatment increased T regulatory cells and reduced the levels of IL6 in the synovium of arthritic mice which were associated with reduced pain. Treated mice showed P. histicola colonization in the duodenum, a niche from where it was isolated. Further, P. histicola treatment showed varied effects based on circadian rhythm. A longitudinal follow-up of the gut microbial profile in different gut sections suggested that dysbiosis caused due to arthritis in DQ8 is partly normalized to naïve profile after treatment with P. histicola. Treated mice showed an expansion of genus Allobaculum, a butyrate-producing Firmicutes in the guts of DQ8 arthritic mice as compared to pre-treatment. The post-treatment restoration was also reflected in short-chain fatty acids and glucose assimilation rates in T cells. The present study proposes that P. histicola treatment of arthritis in DQ8 mice leads to gut homeostasis. Normalized microbial diversity and metabolites lead to immune regulation resulting in reduced inflammation with improved physical function. The observations suggest P. histicola as a strong candidate for monoclonal bio-therapeutics.

Isolation, characterization, and application of thermotolerant Streptomyces sp. K5 for efficient conversion of cellobiose to chitinase using pulse- feeding strategy

In this study, the selection of target sugars in lignocellulosic hydrolysate and the implementation of a bioprocess strategy led to efficient production of a catalyst for chitinous bioresource conversion. Streptomyces sp. K5, a chitinase producer in the presence of glucose, was isolated at 50 °C on an agar plate with glucose and colloidal chitin. The K5 was found to produce chitinase with a maximum activity of 70 U/L in a medium containing glucose and xylose as well as colloidal chitin as an inducer. The assimilation of glucose and xylose, however, was extremely slow, with significant residual being present even after 168 h of incubation. Assimilation tests of glucose, xylose, and cellobiose confirmed that K5 produces chitinase without chitinous inducer and assimilates cellobiose much more rapidly than either glucose or xylose. At the same time, the complete exhaustion of each sugar initiated chitinase inactivation. A pulse- feeding strategy was adopted for the cellobiose, taking its rapid assimilation, β-glucosidase activity, and chitinase inactivation into account, and a maximum chitinase activity of 235 U/L was achieved under pulse- feeding conditions that included four pulses.

Effects of mutations of GID protein-coding genes on malate production and enzyme expression profiles in Saccharomyces cerevisiae.

During alcohol fermentation, Saccharomyces cerevisiae produces organic acids, including succinate, acetate, and malate. Since malate contributes to the pleasant flavor of sake (a Japanese alcoholic beverage), various methods for breeding high-malate-producing yeast have been developed. We previously isolated a high-malate-producing strain and found that a missense mutation in GID4 was responsible for the high-malate-producing phenotype. Gid4 is a component of the GID (glucose-induced degradation-deficient) complex and stimulates the catabolic degradation of gluconeogenic enzymes. In this study, the mechanism by which this mutation led to high malate production in yeast cells was investigated. The evaluation of disruptants and mutants of gluconeogenic enzymes revealed that cytosolic malate dehydrogenase (Mdh2) participated in the malate production. Furthermore, target proteome analysis indicated that an increase in malate production resulted from the accumulation of Mdh2 in gid4 disruptant due to the loss of GID complex-mediated degradation. Next, we investigated the effects of GID protein-coding genes (GID1-GID9) on organic acid production and enzyme expression profiles in yeast. The disruptants of GID1, 2, 3, 4, 5, 8, and 9 exhibited high malate production. Comparison of protein abundance among the GID disruptants revealed variations in protein expression profiles, including in glycolysis and tricarboxylic acid cycle-related enzymes. The high-malate-producing disruptants showed the activation of several glycolytic enzymes and a reduction in enzymes involved in the conversion of pyruvate to ethanol. Our results suggest that high-malate-producing disruptants adapt their metabolism to produce malate in excess via the regulation of protein expression in glucose assimilation and ethanol fermentation. KEY POINTS: An increase in malate level of GID4 mutant resulted from the accumulation of Mdh2. The disruptants of GID1, 2, 3, 4, 5, 8, and 9 showed high malate production. The protein expression profiles in the GID disruptants differed from one another.

Improving glucose and xylose assimilation in Azotobacter vinelandii by adaptive laboratory evolution.

Azotobacter vinelandii is a microorganism with biotechnological potential because its ability to produce alginate and polyhydroxybutyrate. Large-scale biotechnological processes are oriented to sustainable production by using biomass hydrolysates that are mainly composed by glucose and xylose. In the present study, it was observed that A. vinelandii was unable to consume xylose as the sole carbon source and that glucose assimilation in the presence of xylose was negatively affected. Adaptive Laboratory Evolution (ALE) was used as a metabolic engineering tool in A. vinelandii, to improve both carbohydrate assimilation. As a result of ALE process, the CT387 strain was obtained. The evolved strain (CT387) grown in shaken flask cultivations with xylose (8gL-1) and glucose (2gL-1), showed an increase of its specific growth rate (µ), as well as of its glucose and xylose uptake rates of 2, 6.45 and 3.57-fold, respectively, as compared with the parental strain. At bioreactor level, the µ, the glucose consumption rate and the relative expression of gluP that codes for the glucose permease in the evolved strain were also higher than in the native strain (1.53, 1.29 and 18-fold, respectively). Therefore, in the present study, we demonstrated the potential of ALE as a metabolic engineering tool for improving glucose and xylose consumption in A. vinelandii.

Osmotrophic glucose and leucine assimilation and its impact on EPA and DHA content in algae.

The uptake of dissolved organic compounds, that is, osmotrophy, has been shown to be an efficient nutritional strategy for algae. However, this mode of nutrition may affect the biochemical composition, for example, the fatty acid (FA) contents, of algal cells. This study focused on the osmotrophic assimilation of glucose and leucine by selected seven algal strains belonging to chlorophytes, chrysophytes, cryptophytes, dinoflagellates and euglenoids. Our laboratory experiments with stable isotope labeling showed that osmotrophy occurred in four of the selected seven strains. However, only three of these produced long chain omega-3 FAs eicosapentaenoic acid (EPA; 20:5ω3) and docosahexaenoic acid (DHA; 22:6ω3). High glucose content (5 mg L−1) affected negatively on the total FAs of Mallomonas kalinae and the total omega-3 FAs of Cryptomonas sp. Further, glucose assimilation explained 35% (negative effect) and leucine assimilation 48% (positive effect) of the variation of EPA, DHA and the FAs related to their synthesis in Cryptomonas sp. Moderate glucose concentration (2 mg L−1) was found to enhance the growth of Cryptomonas ozolinii, whereas low leucine (20 µg L−1) enhanced the growth of M. kalinae. However, no systematic effect of osmotrophy on growth rates was detected. Our study shows that osmotrophic assimilation of algae is species and compound specific, and that the effects of the assimilated compounds on algal metabolism also varies depending on the species.

Lowering effect of viable Pediococcus pentosaceus QU 19 on the rise in postprandial glucose.

In the present study, we investigated the glucose-decreasing action of lactic acid bacteria (LAB). The finding of this study could be helpful for people in controlling their blood sugar levels. The LAB candidate was isolated from a Japanese fermented food and identified as Pediococcus pentosaceus by an analysis of its genome sequence. Postprandial blood glucose elevation was investigated using oral starch tolerance tests in mice. Normal mice were fed starch and lyophilized cells of P. pentosaceus QU 19 at the same time. Even without pre-administration of P. pentosaceus QU 19, elevation of the blood glucose level was significantly suppressed by the intake of P. pentosaceus QU 19 at the same time as oral administration of starch. According to the results for its survival in simulated digestive juice and the reduction of blood glucose level in mice, P. pentosaceus QU 19 has potential hypoglycemic activity. In vitro measurements revealed that the glucose-decreasing action of P. pentosaceus QU 19 is probably caused by the glucose assimilation of the strain, not the inhibition of carbohydrate-splitting enzymes which has been reported for other LABs previously. These findings indicate that specific strains of LAB, especially P. pentosaceus QU 19, and foods fermented by LAB may be beneficial for people who must manage glucose ingestion.

Identification of a cell-surface protein involved in glucose assimilation and disruption of the crystalline region of cellulose by Cytophaga hutchinsonii.

The crystalline region of cellulose is the main barrier to the utilization of crystalline cellulose. Cytophaga hutchinsonii actively digests the crystalline region of cellulose by an unknown mechanism. Transposon mutagenesis was done to identify a novel gene locus chu_1557, which is required for efficient disruption of the crystalline region of cellulose, and the absence of CHU_1557 resulted in decreased glucose assimilation efficiency. The defect of the mutant in the disruption of the crystalline region of cellulose was partially retained by additional glucose or pre-culturing the mutant in a low glucose concentration medium which could improve its glucose absorption efficiency. These results suggested that extracellular glucose has important roles in the disruption of crystalline cellulose by C. hutchinsonii. Further study showed that the expression of an outer membrane protein CHU_3732 was downregulated by the absence of CHU_1557 in a low glucose concentration medium. CHU_3732 was involved in uptake of glucose and its expression was induced by a low concentration of glucose. CHU_3732 was predicted to be a porin, so we inferred that it may work as a glucose transport channel in the outer membrane. Based on these results, we deduced that CHU_1557 played a role in the process of glucose assimilation and its disruption affected the expression of other proteins related to glucose transportation such as CHU_3732, and then affected the cell growth in a low glucose concentration medium and disruption of the crystalline region of cellulose.

Growth behavior, glucose consumption and phenol removal efficiency of Chlorella vulgaris under the synergistic effects of glucose and phenol

The use of algae is an effective approach to remove phenol and its derivatives from polluted water. The growth behavior, glucose consumption and phenol removal efficiency of Chlorella vulgaris under the synergistic effects of glucose and phenol were investigated. The evolutions of tolerance and removal efficiency of C. vulgaris to phenol under different trophic modes and glucose contents were observed. The results revealed that growth of C. vulgaris were inhibited with the increase of phenol from 0 to 400 mg L−1 in culture media; the tolerance to phenol enhanced with the addition of glucose from 2 to 10 g L−1, while glucose consumption was inhibited with the increase of phenol content; phenol removal efficiency varied with glucose concentrations in mixotrophic media. The finding suggested that phenol inhibited the growth of C. vulgaris and glucose assimilation under mixotrophic cultivation, while appropriate glucose addition could enhance the tolerance of C. vulgaris to phenol and affect the phenol removal efficiency.

Investigating the modulation of metabolites under high light in mixotrophic alga Asteracys sp. using a metabolomic approach

Metabolomic fluctuations in microalgal cells under mixotrophic growth regimes are scarcely understood. Our study explores these changes in an oleaginous microalga, Asteracys sp. when grown mixotrophically under high irradiances. Biomass production under autotrophic and mixotrophic regimes were studied at two light intensities (LL; 100 μmol photons m−2 s−1 and HL; 900 μmol photons m−2 s−1) for Asteracys sp. Studies on supplementation of external carbon such as glucose (mixotrophic mode) under different light intensities has not been demonstrated in context with enhanced biomass production. In the present study, we have provided external source of carbon as glucose in LL and HL regimes and observed a marked influence of glucose assimilation on the overall machinery of the cell. Our qualitative metabolomics reveals a stress-like phenomenon in Asteracys sp. under HL with enhanced biomass. In overview, our data analysis highlights few metabolites such as trehalose, proline and α-tocopherol that showed significant changes in response to light as well as in growth regime. Our analysis predicts an interrelation cross talk between maltose and sucrose pathways within cells owing to possible alteration in starch degradation under the influence of light. However, mixotrophy with additional carbon supplementation in presence of light did not alter or enhance lipid biosynthesis. In conclusion, our hypothesis indicates modulation of metabolites in presence of glucose under high light would rather facilitate rearrangement in metabolic pathways for enhanced biomass production.

Light irradiation can regulate the growth characteristics and metabolites compositions of Rhodotorula mucilaginosa.

Light is an important factor that can induce the growth of varieties of organisms including fungi and their secondary metabolites. The evolutions of biomass, carotenoids, lipid production, compositions and contents of fatty acid and amino acid in Rhodotorula mucilaginosa were investigated under different light irradiation conditions. The results indicated that irradiation with 1700lx could promote the growth and glucose assimilation of R. mucilaginosa, compared to the dark control, while the trial with 3500lx had certain inhibiting effects. The carotenoids concentrations and percentages of unsaturated fatty acid (USFA, C16:1 and C18:1) increased with the improvement of irradiation intensity. Conversely, the proportions of saturated fatty acids (C16:0, C18:0 and C20:0) were decreased. The relative contents of amino acid and total protein were reduced under illumination compared to dark control. Conclusively, irradiation could change the cell growth and metabolites of the pigmented fungus, which implied that there may be a photoinduced mode exists in R. mucilaginosa similar to that of Neurospora crassa, and it also could be applied to regulate the biosynthesis and production of valuable components such as carotenoids and USFA.

Functional implementation of a linear glycolysis for sugar catabolism in Pseudomonas putida

The core metabolism for glucose assimilation of the soil bacterium and platform strain Pseudomonas putida KT2440 has been reshaped from the native, cyclically-operating Entner-Doudoroff (ED) pathway to a linear Embden-Meyerhof-Parnas (EMP) glycolysis. The genetic strategy deployed to obtain a suitable host for the synthetic EMP route involved not only eliminating enzymatic activities of the ED pathway, but also erasing peripheral reactions for glucose oxidation that divert carbon skeletons into the formation of organic acids in the periplasm. Heterologous glycolytic enzymes, recruited from Escherichia coli, were genetically knocked-in in the mutant strain to fill the metabolic gaps for the complete metabolism of glucose to pyruvate through a synthetic EMP route. A suite of genetic, physiological, and biochemical tests in the thereby-refactored P. putida strain—which grew on glucose as the sole carbon and energy source—demonstrated the functional replacement of the native sugar metabolism by a synthetic catabolism. 13C-labelling experiments indicated that the bulk of pyruvate in the resulting strain was generated through the metabolic device grafted in P. putida. Strains carrying the synthetic glycolysis were further engineered for carotenoid synthesis from glucose, indicating that the implanted EMP route enabled higher carotenoid content on biomass and yield on sugar as compared with strains running the native hexose catabolism. Taken together, our results highlight how conserved metabolic features in a platform bacterium can be rationally reshaped for enhancing physiological traits of interest.

Expression of genes related to glucose metabolism and joint destruction in the development of diabetes mellitus in patients with osteoarthritis

Many patients with osteoarthritis (OA) tend to have comorbidities. This tendency is more frequently observed to increase with age. One of the comorbidities is type 2 diabetes mellitus (T2DM). Due to the higher prevalence of coexistence of these two conditions, it has been suggested that T2DM-associated hyperglycemia may adversely affect joint tissues and increase OA severity. However, the molecular mechanisms in the development of DM in patients with OA remain unclear. Objective : to trace the dynamics of T2DM development in patients with OA at the level of the expression of genes associated with glucose metabolism, joint destruction, and general regulation of metabolic processes. Patients and methods . Three patients with OA were followed up for 4—6 years, including the year of onset of T2DM. The clinical condition of the patients was analyzed once a year. Total RNA was annually isolated from their blood and used to determine the level of gene expression by real-time polymerase chain reaction. Results and discussion. The development of T2DM was shown to be accompanied by the increased expression of genes related to glycolysis, Krebs cycle, pentose phosphate pathway, matrix metalloproteinases and regulators of AMPKand mTOR metabolism. By contrast, the level of the hypoxia regulator HIF1a and hexosamine pathway genes was decreased. Conclusion . The occurrence of T2DM in the presence of OA is likely to be associated with the higher needs for cells for ATP energy and is accompanied by activation of the glucose assimilation pathways, as well as by the increased expression of the genes responsible for extracellular matrix destruction. This may be caused by impaired protein glycosylation due to inhibition of the hexosamine pathway.

Влияние источника углерода на синтез биомассы и экзополисахаридов бактериями Paenibacillus mucilaginosus

Paenibacillus mucilaginosus bacteria are one of the most promising soil microorganisms for obtaining microbiological fertilisers. These bacteria produce indolyl-3-acetous acid and ammonia, as well as possessing the capability of nitrogen fixation and solubilisation of undissolved soil minerals yielding potassium and phosphorus accessible to plants and consequently stimulating plant growth. P. mucilaginosus biological preparations are also useful in the form of fertiliser additives leading to immune stimulation and increase in the productivity of agricultural animals. For the cultivation of P. mucilaginosus bacterial strains, sucrose is recommended. At the same time, little research has been carried out on the influence of glucose and fructose on the growth of these bacteria. The aim of the work was to study the effect of various carbon sources in the composition of the nutrient medium on the biomass synthesis of P. mucilaginosus strains. As a result, a double growth cycle was observed under the conditions of culture on a nutrient medium with a sucrose and a glucose-fructose mixture. The specific growth rate of the bacterial strains under consideration in the assimilation of glucose is higher than in the assimilation of fructose. Accordingly, the generation time for glucose assimilation turned out to be lower than that for fructose assimilation. The growth of the considered bacterial strains on a single fructose nutrient medium was not observed. Glucose demonstrated similar effectiveness of application in culture media to sucrose for the cultivation of all considered bacterial strains of P. mucilaginosus. During culturing of P. mucilaginosus bacteria on nutrient media from glucose and sucrose, the yield of biomass and exopolysaccharides was established to be higher than for the case of a nutrient medium presented by a glucose-fructose mixture. Promising strains for industrial application include 563, 567 and 574, providing a high yield of biomass and exopolysaccharides with culture on a glucose-admixed nutrient medium, and 560, 568 and 572 strains when cultivated on a nutrient medium containing sucrose.