Mixed Sugars Research Articles

A method for preparing N-alkylated kanosamines from diacetone d-glucose

The aminoglycoside (AG) antibiotics have seen a resurgence in their clinical use given the increase in multi drug resistant bacterial infections. Campaigns to generate novel analogs show promise that structural modification can lead to compounds with improved pharmacological properties. The results described herein include a new method to synthesize mono-, di-, and mixed N-alkylated kanosamine sugars and their elaboration into novel glycosides that inhibit bacterial protein synthesis in vitro.

PKA and HOG signaling contribute separable roles to anaerobic xylose fermentation in yeast engineered for biofuel production.

Lignocellulosic biomass offers a sustainable source for biofuel production that does not compete with food-based cropping systems. Importantly, two critical bottlenecks prevent economic adoption: many industrially relevant microorganisms cannot ferment pentose sugars prevalent in lignocellulosic medium, leaving a significant amount of carbon unutilized. Furthermore, chemical biomass pretreatment required to release fermentable sugars generates a variety of toxins, which inhibit microbial growth and metabolism, specifically limiting pentose utilization in engineered strains. Here we dissected genetic determinants of anaerobic xylose fermentation and stress tolerance in chemically pretreated corn stover biomass, called hydrolysate. We previously revealed that loss-of-function mutations in the stress-responsive MAP kinase HOG1 and negative regulator of the RAS/Protein Kinase A (PKA) pathway, IRA2, enhances anaerobic xylose fermentation. However, these mutations likely reduce cells’ ability to tolerate the toxins present in lignocellulosic hydrolysate, making the strain especially vulnerable to it. We tested the contributions of Hog1 and PKA signaling via IRA2 or PKA negative regulatory subunit BCY1 to metabolism, growth, and stress tolerance in corn stover hydrolysate and laboratory medium with mixed sugars. We found mutations causing upregulated PKA activity increase growth rate and glucose consumption in various media but do not have a specific impact on xylose fermentation. In contrast, mutation of HOG1 specifically increased xylose usage. We hypothesized improving stress tolerance would enhance the rate of xylose consumption in hydrolysate. Surprisingly, increasing stress tolerance did not augment xylose fermentation in lignocellulosic medium in this strain background, suggesting other mechanisms besides cellular stress limit this strain’s ability for anaerobic xylose fermentation in hydrolysate.

Efficient accumulation of sclerotiorin via overcoming low pH caused by overflow carbon metabolism during cell suspension culture of Penicillium sclerotiorum

Abstract Sclerotiorin is the secondary metabolite of Penicillium sclerotiorum, which is usually produced by solid state fermentation or liquid culture without stirring (formation of mycelial mat on air-liquid surface). In the present work, it was confirmed that malic acid was produced during cell suspension culture in a liquid medium (mycelia acting as planktonic cells) using resting cells as whole cell biocatalyst. The accumulation of malic acid was caused by overflow carbon metabolism at high glucose concentration (Crabtree effect), which led to low ambient pH value. The low ambient pH repressed sclerotiorin biosynthesis. In order to overcome the low pH caused by Crabtree effect, many influenced factors, such as phosphate buffer system, initial pH, biomass loading, slowly metabolized carbon resource, and mixed sugars as carbon resource, were investigated. Finally, the low pH caused by Crabtree effect was eliminated by taking the advantage of carbon metabolite repression via utilization of mixed sugars (glucose and lactose) as carbon resource. And then, an ideal pH approximately 3.8 was maintained and corresponding high concentration of sclerotiorin (150 absorbance unit at 365 nm, corresponding to 1.9 g/l) was achieved by cell suspension culture with a relatively high biomass loading 11.7 g/l DCW (dry cell weight).

Antioxidant activity changes of exopolysaccharides with different carbon sources from Lactobacillus plantarum LPC-1 and its metabolomic analysis.

The effects of different carbon sources on the antioxidant activity changes of exopolysaccharides (EPSs) were determined for the strains Lactobacillus plantarum LPC-1 with glucose, sucrose and its mixture as carbon sources, respectively. Meanwhile, GC-MS datasets coupled with multivariate statistical methods were used to investigate metabolic changes of EPSs-producing L. plantarum cultured with different carbon source. Among carbon sources examined, both of glucose and sucrose were favorable for the cell growth, while the maximum EPSs yield was achieved when sucrose was employed. EPSs cultured with different carbon sources showed remarkable different antioxidant activities, and EPSs with sucrose or mixed sugar as carbon source exhibited a promising antioxidant activity, such as hydroxyl scavenging activity and DPPH radical scavenging activity. Results from rice cultivation showed a similar conclusion that there were also significant differences in the antioxidant activities of EPSs obtained from different carbon sources in inducing rice resistance to chromium stress, but addition of EPSs had no significant impact on the uptake of Cr metals. Principal component analysis showed clear differences in metabolites among different treatment, and the glycolysis and tricarboxylic acid cycle were decreased when sucrose or mixed sugar was used as carbon source, and the production of lactic acid was also reduced, which might be the main reasons for the overproduction of EPSs. Our results indicated that Lactobacillus strain, depending on the carbon source in the medium, could produce EPSs of different biological properties, and the metabolomic analysis findings provided the first omics view of cell growth and EPSs synthesis in L. plantarum, which would be a theoretical basis for further improving the production of EPSs.

Kinetic study of butanol production from mixtures of glucose and xylose and investigation of different pre-growth strategies

This study proposes a dynamic model that describes key characteristics of fermentative butanol production from glucose and xylose mixtures. The model has 12 parameters and incorporates noncompetitive inhibitory interaction between sugars as well as inhibitions due to high substrate and butanol concentrations. Different pre-growth strategies to achieve co-utilization of sugars were explored together with their effects on fermentation kinetics. Mixed sugar fermentation by the cultures pre-grown on a mixture of glucose and xylose showed a higher endurance to inhibition, a 2-fold increase in butanol production and a 1.5-fold increase in total sugar consumption compared to cultures pre-grown on xylose only. The average squared correlation coefficients (r2) between experimental observations and model predictions were 0.917 and 0.926 for fermentations done by the cultures pre-grown on xylose only, and pre-grown on a mixture of glucose and xylose, respectively. Sensitivity analysis on the model parameters revealed that the growth parameters were the most critical. The proposed model can serve as a basis for modeling of microbial butanol production from lignocellulosic biomass and be applied to other substrates and microorganisms.

Karakteristik Nilai Kalor Briket Campuran Tebu Tibarau dan Kulit Durian

Kebutuhan energi yang meningkat di masyarakat, dapat berdampak terhadap kelangkaan bahan bakar minyak sehingga menjadi ketergantungan terhadap energi sebagai kebutuhan utama. Pengembangan diversifikasi produk berupa briket campuran tebu tibarau dan kulit durian sebagai upaya dalam mendapatkan bahan bakar alternatif. Sumber energi terbarukan yang berpotensi dijadikan bahan bakar briket berbahan baku tebu tibarau dan kulit durian. Upaya dan inovasi briket campuran tebu tibarau dan kulit durian yang dihasilkan sebagai bahan bakar alternatif sehingga mempunyai karakteristik terhadap kualitas dan performa yaitu nilai kalornya. Tahapan ujicoba dan evaluasi terhadap briket campuran tebu tibarau dan kulit durian diperlukan sebelum dikembangkan dan direkomendasi sebagai bahan bakar alternatif. Melalui teknologi proses pembuatan produk briket dengan kompaksi dan optimalisasi kandungan komposisi dan penggunaan jenis perekat sehingga diperoleh nilai kalornya. Dari penelitian ini diperoleh briket campuran tebu tibarau dan kulit durian yang direkomendasikan sebagai bahan bakar alternatif. Karakteristik nilai kalor briket campuran tebu tibarau dan kulit durian yang tertinggi dengan perekat gambir diperoleh sebesar 16967,91 kJ/Kg dan massa jenis 682,23 Kg/m3. Briket (biomassa) yang memiliki massa jenis (densitas) yang tinggi memiliki nilai kalor yang tinggi pula. Perbandingan komposisi campuran pada pembuatan briket yaitu 40% : 40% : 20%. Komposisi campuran antara bahan baku dengan perekat, besaran butir, gaya penekanan (kompaksi) sangat mempengaruhi karakteristik nilai kalor dan massa jenis briket tersebut.

Highly efficient continuous acetone–butanol–ethanol production from mixed sugars without carbon catabolite repression

Factors including the composition and concentration of mixed sugars, dilution rate, and cell density were examined to establish a highly efficient continuous acetone–butanol–ethanol (ABE) fermentation process using Clostridium saccharoperbutylacetonicum N1-4. During conventional continuous fermentation at a dilution rate of 0.15 h−1, the glucose/xylose mixture exhibited carbon catabolite repression (CCR) with an intensity (ICCR) of 4.18 and ABE productivity of 0.574 g L−1 h−1, while CCR was alleviated using a cellobiose/xylose mixture with 1.60 and 1.09 g L−1 h−1, respectively. Continuous fermentation with high cell density by cell recycling improved ABE productivity to 5.32 g L−1 h−1 at a dilution rate of 0.686 h−1 owing to high xylose consumption. Further optimization of mixed sugar concentration in the feeding medium induced the best performance of continuous ABE production from mixed sugar without CCR, relative to previous studies: ABE productivity, 6.09 g L−1 h−1; ABE yield, 0.214 g g−1; ICCR, 1.48.

Co-culture strategy for improved 2G bioethanol production using a mixture of sugarcane molasses and bagasse hydrolysate as substrate

Mixed sugar co-fermentation represents an interesting fermentation strategy to improve bioethanol production from low-cost raw materials, such as sugarcane bagasse. The aim of this study was to evaluate a mixture of sugars (hexoses and pentoses) from sugarcane bagasse hemicellulosic hydrolysate diluted with sugarcane molasses as substrate to be converted either in monoculture or co-culture fermentations. A co-culture fermentation technique using hexoses-fermenting yeasts (Saccharomyces cerevisiae: from Santa Adélia Mill, an industrial strain (SA) and a respiratory-deficient mutant strain (ScP)) together with xylose-fermenting yeasts (Scheffersomyces stipitis (SS) and Spathaspora passalidarum (SP)) was carried out to enhance bioethanol production. Co-culture fermentation in a medium with a high mixed sugar concentration (100 g.L−1) resulted in a complete sugar consumption, ethanol titer of 30.2 g.L−1 and increased ethanol productivity (Qp =4.44 g.L−1. h−1) using wild type strains (SP + SA). In contrast, the maximum ethanol titer (49.2 g.L−1) was reached during co-culture performed with S. stipitis and the respiratory-deficient mutant strain of S. cerevisiae, however ethanol productivity (Qp =0.60 g.L−1. h−1) decreased due to the poor fermentation performance of the mutant strain. This work suggests that improved efficiency and good scalability of the mixed sugar fermentation process makes an important basis for the economic viability of bioethanol production from lignocelluloses.

Improving co-fermentation of glucose and xylose by adaptive evolution of engineering xylose-fermenting Saccharomyces cerevisiae and different fermentation strategies

Xylose utilization of engineered yeast is vulnerable to inhibitors generated during pretreatment of lignocellulose. In this study, adaptive evolution was applied to enhance the tolerance of xylose-fermenting strain. Compared to the parental strain, the ethanol yield was increased by 60% and 80% for the adapted strain (E7-403) when xylose was used as the sole carbon resource with 20% and 50% inhibitor cocktails, respectively. E7-403 removed furfural more effectively than parental strain (E7) in the fermentation with 100% inhibitor cocktails. In the fermentation with mixed sugar and high inhibitor concentration, glucose was depleted within 36 h for E7-403 while 6.1 g/L glucose was still left after 120 h for E7. Consequently, ethanol yield of E7-403 was 22.9% higher than that of E7. It was demonstrated that E7-403 strain exhibited an enhanced ability for regulating cellular reactive oxygen species, which alleviated the harmful effects of inhibitors. Meanwhile, E7-403 strain was further applied in co-culture and pre-fermentation process to improve xylose utilization.

Co-culture of Zymomonas mobilis and Scheffersomyces stipitis immobilized in polymeric membranes for fermentation of glucose and xylose to ethanol

Submerged hollow fiber membrane bioreactor (SHFMB) was fabricated and operated to co-culture Zymomonas mobilis and Scheffersomyces stipitis for fermentation of glucose and xylose mixture. Suspended Z. mobilis and S. stipitis in co-culture exhibited poor xylose fermentation due to substrate/product inhibition and catabolite repression. Immobilization in polymeric membranes was effective in alleviating these inhibitions in the SHFMB, which resulted in concomitant fermentation of glucose and xylose at initial glucose and xylose concentrations of 20–80 g L−1 and 10–40 g L−1, respectively. Selective aeration in SHFMB resulted in 100% glucose and >70% xylose fermentation within 48 h, producing 36.7 g L−1 ethanol from 90 g L−1 sugar mixture. Partial immobilization in the membranes allowed Z. mobilis to diffuse in suspension under benign condition, whereas S. stipitis remained inside membrane pores throughout the operation. Xylose fermentation in the SHFMB could be enhanced further by increasing the number of S. stipitis immobilized membranes. Cell immobilization in the SHFMB was stable during 12 consecutive batch runs over 24 days, and exhibited improved xylose fermentation due to gradual increase in S. stipitis accumulation within the membranes. These results indicate that SHFMB can be promising for co-culture of Z. mobilis and S. stipitis in fermentation of mixed sugars from lignocellulosic hydrolysate.

Enhancing the Co-utilization of Biomass-Derived Mixed Sugars by Yeasts.

Plant biomass is a promising carbon source for producing value-added chemicals, including transportation biofuels, polymer precursors, and various additives. Most engineered microbial hosts and a select group of wild-type species can metabolize mixed sugars including oligosaccharides, hexoses, and pentoses that are hydrolyzed from plant biomass. However, most of these microorganisms consume glucose preferentially to non-glucose sugars through mechanisms generally defined as carbon catabolite repression. The current lack of simultaneous mixed-sugar utilization limits achievable titers, yields, and productivities. Therefore, the development of microbial platforms capable of fermenting mixed sugars simultaneously from biomass hydrolysates is essential for economical industry-scale production, particularly for compounds with marginal profits. This review aims to summarize recent discoveries and breakthroughs in the engineering of yeast cell factories for improved mixed-sugar co-utilization based on various metabolic engineering approaches. Emphasis is placed on enhanced non-glucose utilization, discovery of novel sugar transporters free from glucose repression, native xylose-utilizing microbes, consolidated bioprocessing (CBP), improved cellulase secretion, and creation of microbial consortia for improving mixed-sugar utilization. Perspectives on the future development of biorenewables industry are provided in the end.

Elegant and Efficient Biotransformation for Dual Production of d-Tagatose and Bioethanol from Cheese Whey Powder.

In this study, the dual production of valuable d-tagatose and bioethanol from lactose and cheese whey powder is presented. First, a one-pot biosynthesis involving lactose hydrolysis and d-galactose isomerization for d-tagatose production was established using crude enzymes of recombinant Escherichia coli with l-arabinose isomerase (L-AI) at 50 °C. Compared to the current enzymatic system, only L-AI was overexpressed, because of the unexpectedly thermotolerant β-galactosidase in E. coli BL21(DE3). Moreover, this high temperature rendered the d-glucose catabolism of E. coli inactive, while retaining all fermentable sugars for bioethanol fermentation. Thereafter, the mixed sugar syrup was fermented by Saccharomyces cerevisiae NL22. A total of 23.5 g/L d-tagatose and 26.9 g/L bioethanol was achieved from cheese whey powder containing 100 g/L lactose. This bioprocess not only provides an efficient method for the functionalization of byproduct whey, but also offsets the high production cost of d-tagatose and bioethanol.

Integration of Pretreatment With Simultaneous Counter-Current Extraction of Energy Sorghum for High-Titer Mixed Sugar Production

Sorghum (Sorghum bicolor L. Moench) offers substantial potential as a feedstock for the production of sugar-derived biofuels and biochemical products from cell wall polysaccharides (i.e., cellulose and hemicelluloses) and water-extractable sugars (i.e., glucose, fructose, sucrose, and starch). A number of preprocessing schemes can be envisioned that involve processes such as sugar extraction, pretreatment, and densification that could be employed in decentralized, regional-scale biomass processing depots. In this work, an energy sorghum exhibiting a combination of high biomass productivity and high sugar accumulation was evaluated for its potential for integration into several potential biomass preprocessing schemes. This included counter-current extraction of water-soluble sugars followed by mild NaOH or liquid hot water pretreatment of the extracted bagasse. A novel processing scheme was investigated that could integrate with current diffuser-type extraction systems for sugar extraction. In this approach, mild NaOH pretreatment (i.e., 8%) at high metabolic yields without detoxification using a Saccharomyces cerevisiae strain metabolically engineered and evolved to ferment xylose.

Optimized xylose isomerase uptake and expression level in Saccharomyces cerevisiae for improving ethanol production

The ability to engineer the yeast Saccharomyces cerevisiae to efficiently convert lignocellulosic biomass to ethanol remains a considerable challenge. Here, we propose a new reprogrammable strategy to optimize the expression level of the xylose isomerase (XI) gene with the induction of mutations in S . cerevisiae to improve efficient ethanol production and productivity. We sought to fine-tune the xylose uptake and catabolism abilities of S. cerevisiae during fermentation by improving efficiency of the xylose transporter, which was fused with four copies of the XI gene under the control of different promoters to obtain recombinant yeast strains. In fermentation experiments, the optimized strain CW9 cultured in yeast extract-peptone (YP) medium containing approximately 65 g/L glucose and 55 g/L xylose produced consistent ethanol yields of 0.45 g/g total sugar in about 72 h, which was close to 90% of the theoretical yield. These promising results indicate that strain CW9 is the best producer of ethanol from mixed sugar when synthetically regulating the xylose assimilation pathway. Overall, this study provides an optimal method to control XI expression levels to find better conditions for enhancing biofuel production.

Evolutionary engineered Candida intermedia exhibits improved xylose utilization and robustness to lignocellulose-derived inhibitors and ethanol

The development of robust microorganisms that can efficiently ferment both glucose and xylose represents one of the major challenges in achieving a cost-effective lignocellulosic bioethanol production. Candida intermedia is a non-conventional, xylose-utilizing yeast species with a high-capacity xylose transport system. The natural ability of C. intermedia to produce ethanol from xylose makes it attractive as a non-GMO alternative for lignocellulosic biomass conversion in biorefineries. We have evaluated the fermentation capacity and the tolerance to lignocellulose-derived inhibitors and the end product, ethanol, of the C. intermedia strain CBS 141442 isolated from steam-exploded wheat straw hydrolysate. In a mixed sugar fermentation medium, C. intermedia CBS 141442 co-fermented glucose and xylose, although with a preference for glucose over xylose. The strain was clearly more sensitive to inhibitors and ethanol when consuming xylose than glucose. C. intermedia CBS 141442 was also subjected to evolutionary engineering with the aim of increasing its tolerance to inhibitors and ethanol, and thus improving its fermentation capacity under harsh conditions. The resulting evolved population was able to ferment a 50% (v/v) steam-exploded wheat straw hydrolysate (which was completely inhibitory to the parental strain), improving the sugar consumption and the final ethanol concentration. The evolved population also exhibited a better tolerance to ethanol when growing in a xylose medium supplemented with 35.5 g/L ethanol. These results highlight the potential of C. intermedia CBS 141442 to become a robust yeast for the conversion of lignocellulose to ethanol.

Engineering Saccharomyces cerevisiae for Enhanced Production of Protopanaxadiol with Cofermentation of Glucose and Xylose.

Protopanaxadiol (PPD), an active triterpene compound, is the precursor of high-value ginsenosides. In this study, we report a strategy for the enhancement of PPD production in Saccharomyces cerevisiae by cofermentation of glucose and xylose. In mixed sugar fermentation, strain GW6 showed higher PPD titer and yield than that obtained from glucose cultivation. Then, engineering strategies were implemented on GW6 to enhance the PPD yields, such as adjustment of the central carbon metabolism, optimization of the mevalonate pathway, reinforcement of the xylose assimilation pathway, and regulation of cofactor balance, namely, overexpression of xPK/PTA, ERG10/ERG12/ERG13, XYL1/XYL2/TAL1, and POS5, respectively. In particular, the final obtained strain GW10, harboring overexpressed POS5, exhibited the highest PPD yield, which was 2.06 mg of PPD/g of mixed sugar. In a 5-L fermenter, PPD titer reached 152.37 mg/L. These promising results demonstrate the great advantages of mixed sugar over glucose for high-yield production of PPD.

Calculating Intake of Dietary Risk Components Used in the Global Burden of Disease Studies from the What We Eat in America/National Health and Nutrition Examination Surveys.

Disability adjusted life years (DALYs) is a health burden metric that combines years of life lost due to disease disability and premature mortality. The Global Burden of Disease (GBD) has been using DALYs to determine the health burden associated with numerous health risks, including risks associated with dietary intakes, at the global and national level. To translate such information at the food level in the U.S., variables in What We Eat in America (WWEIA) need to be aligned with those in the GBD. In this paper, we develop the necessary new variables needed to account for differences in definitions and units between WWEIA and the GBD. We use the Food Patterns Equivalents Database, Food Patterns Equivalents Ingredient Database, Food and Nutrient Database for Dietary Studies, and Standard Reference databases that provide data for WWEIA to develop food group and nutrient variables that align with definitions and units used in the GBD. Considerable effort was needed to disaggregate mixed dishes to GBD components. We also developed a new “non-starchy” vegetable variable, since the GBD vegetables do not include potatoes and corn, and we report fruits and vegetables in grams instead of household measures. New fiber variables were created to avoid double counting of fiber from legumes, whole grains, fruits, and vegetables. Regression analyses were used to predict trans-fat content for foods in WWEIA with missing or incomplete information. The majority of foods in various U.S. Department of Agriculture (USDA) categories contain multiple GBD food groups (e.g., vegetables, whole grains, and processed meat). For most nutrients considered in the GBD, composition is more evenly distributed across the main food categories; however, seafood omega-3 fats were predominantly from either protein foods or mixed dishes and sugar sweetened beverages were from a single category. Dietary intakes in the U.S. fall short of recommendations for all food groups/nutrients with established theoretical minimum-risk targets in GBD. To our knowledge, this is the first approach that aligns WWEIA intake variables with those used in the health burden-based GBD reports. These methods will facilitate researchers to begin comparing data from the U.S. with that from other countries, as well as assess food sustainability performances by concomitantly evaluating DALYs for environmental and nutritional impacts.

Augmenting Pentose Utilization and Ethanol Production of Native Saccharomyces cerevisiae LN Using Medium Engineering and Response Surface Methodology.

Economics of ethanol production from lignocellulosic biomass depends on complete utilization of constituent carbohydrates and efficient fermentation of mixed sugars present in biomass hydrolysates. Saccharomyces cerevisiae, the commercial strain for ethanol production uses only glucose while pentoses remain unused. Recombinant strains capable of utilizing pentoses have been engineered but with limited success. Recently, presence of endogenous pentose assimilation pathway in S. cerevisiae was reported. On the contrary, evolutionary engineering of native xylose assimilating strains is promising approach. In this study, a native strain S. cerevisiae LN, isolated from fruit juice, was found to be capable of xylose assimilation and mixed sugar fermentation. Upon supplementation with yeast extract and peptone, glucose (10%) fermentation efficiency was 78% with ~90% sugar consumption. Medium engineering augmented mixed sugars (5% glucose + 5% xylose) fermentation efficiency to ~50 and 1.6% ethanol yield was obtained with concomitant sugar consumption ~60%. Statistical optimization of input variables Glucose (5.36%), Xylose (3.30%), YE (0.36%), and peptone (0.25%) with Response surface methodology led to improved sugar consumption (74.33%) and 2.36% ethanol within 84 h. Specific activities of Xylose Reductase and Xylitol Dehydrogenase exhibited by S. cerevisiae LN were relatively low. Their ratio indicated metabolism diverted toward ethanol than xylitol and other byproducts. Strain was tolerant to concentrations of HMF, furfural and acetic acid commonly encountered in biomass hydrolysates. Thus, genetic setup for xylose assimilation in S. cerevisiae LN is not merely artifact of xylose metabolizing pathway and can be augmented by adaptive evolution. This strain showed potential for commercial exploitation.

Enhancement of sucrose metabolism in Clostridium saccharoperbutylacetonicum N1-4 through metabolic engineering for improved acetone–butanol–ethanol (ABE) fermentation

This work investigated sucrose metabolism in C. saccharoperbutylacetonicum. Inactivation of sucrose catabolism operon resulted in 28.9% decrease in sucrose consumption and 44.1% decrease in ABE production with sucrose as sole carbon source. Interestingly, a large amount of colloid-like polysaccharides were generated in the mutant, which might be due to inefficient intracellular sucrose metabolism. Deletion of transcriptional repressor gene successfully alleviated CCR and enhanced ABE production by 24.7%. Additional overexpression of endogenous sucrose pathway further elevated sucrose consumption and enhanced ABE production by 17.2%, 45.7%, or 22.5% compared to wild type with sucrose, mixed sugars or sugarcane juice as substrate, respectively. The engineered strain could be a robust platform for efficient biofuel production from inexpensive sucrose-based carbon sources.

Enhanced production of d-lactate from mixed sugars in Corynebacterium glutamicum by overexpression of glycolytic genes encoding phosphofructokinase and triosephosphate isomerase

The use of mixed sugars containing glucose and xylose in lignocellulosic biomass is desirable for the microbial production of chemicals and fuels. We investigated the effect of individual or simultaneous overexpression of glycolytic genes on d-lactate production from a mixture of glucose and xylose by a recombinant xylose-assimilating Corynebacterium glutamicum strain. The individual overexpression of genes encoding phosphofructokinase (PFK) and triosephosphate isomerase (TPI) increased d-lactate production rate by 71% and 34%, respectively, with corresponding increases (2.4- and 1.8-fold) in the glucose consumption; however, the amount of xylose consumed not altered. d-Lactate yield was also increased by 5.5%, but only in the strain overexpressing the gene encoding PFK. In the parent strain and the strains overexpressing the genes encoding PFK or TPI, a reduction in d-lactate production occurred at approximately 900mM after 32h. However, the strain that simultaneously overexpressed the genes encoding PFK and TPI continued to produce d-lactate after 32h, with the eventual production of 1326mM after production for 80h in mineral salts medium. Our findings contribute to the cost-effective, large-scale production of d-lactate from mixed sugars.