Showing papers by "Byung-Gee Kim published in 2017"

Development of High Performance Polyurethane Elastomers Using Vanillin-Based Green Polyol Chain Extender Originating from Lignocellulosic Biomass

Abstract: Vanillin can be obtained from waste of lignocellulosic bioresources with various methods.1−3 Such vanillin was used as chain extender [divanillin-ethanol amine conjugate (DV-EA)] after its dimerization and further modification with ethanolamine in the synthesis of biobased polyurethane, thereby increasing wt % of biocontents in the final polymer. 1,4-Butanediol often used as a general chain extender in polyurethane synthesis was replaced partially with DV-EA. The generated polyurethane hard segment consists of DV-EA polyol and MDI (methylene diisocyanate) units or 1,4-butanediol and MDI units, respectively. The properties of the DV-EA-based polyurethane were investigated with differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), dynamic mechanical analyzer (DMA), X-ray diffraction spectroscopy (XRD), and universal testing machine (UTM). The results showed that this advanced polyurethane has 128% of Young’s modulus and 147% of increased strain compared to those of control, while its st...

Biosynthesis of (-)-5-hydroxy-equol and 5-hydroxy-dehydroequol from soy isoflavone, genistein using microbial whole cell bioconversion

Abstract: Equols are isoflavandiols formed by reduction of soy isoflavones such as daidzein and genistein by gut microorganisms. These phytoestrogens are of interest for their various biological effects. We report biosynthesis from genistein to (−)-5-hydroxy-equol in recombinant E. coli expressing three reductases (daidzein reductase DZNR, dihidrodaidzein reductase DHDR, tetrahydrodaidzein reductase THDR) and a racemase (dihydrodaidzein racemase, DDRC) originating from the gut bacterium, Slackia isoflavoniconvertens. The biosynthesized 5-hydroxy-equol proved as an optically negative enantiomer, nonetheless it displayed an inverse circular dichroism spectrum to (S)-equol. Compartmentalized expression of DZNR and DDRC in one E. coli strain and DHDR and THDR in another increased the yield to 230 mg/L and the productivity to 38 mg/L/h. If the last reductase was missing, the intermediate spontaneously dehydrated to 5-hydroxy-dehydroequol in up to 99 mg/L yield. This novel isoflavene, previously not known to be synthesiz...

Simultaneously Enhancing the Stability and Catalytic Activity of Multimeric Lysine Decarboxylase CadA by Engineering Interface Regions for Enzymatic Production of Cadaverine at High Concentration of Lysine.

Abstract: Cadaverine (1,5-diaminopentane) is a major source of many industrial polyamides such as nylon and chelating agents. Currently, cadaverine is produced by the microbial fermentation of glucose to lysine, which is then decarboxylated by lysine decarboxylase (CadA). However, utilizing CadA for cadaverine production causes enzyme instability. In order to stabilize the CadA homo-decamer structure for in vitro decarboxylation reaction, mutants are designed. Of the four disulfide bond mutants in the multimeric interfacial region, B1 (F14C/K44C) showed a 216-folds increase in the half-life of CadA at 60 °C. On top of B1, another round of mutant screening is performed around F14C and K44C to generate B1/L7M/N8G, which is then examined for cadaverine production (2M lysine and 10% v/v of cell-extract at 50 °C). The reaction pH increased from 4.9 to 8.3, and the final titer of the mutant is 157 g L-1 , that is, 76.7% conversion yield in 9.5 h, whereas the wild-type gave 119 g L-1 , that is, 58.2% conversion yield in 9.5 h.

A Novel Approach for Gene Expression Optimization through Native Promoter and 5′ UTR Combinations Based on RNA-seq, Ribo-seq, and TSS-seq of Streptomyces coelicolor

Abstract: Streptomycetes are Gram-positive mycelial bacteria, which synthesize a wide range of natural products including over two-thirds of the currently available antibiotics. However, metabolic engineering in Streptomyces species to overproduce a vast of natural products are hampered by a limited number of genetic tools. Here, two promoters and four 5′ UTR sequences showing constant strengths were selected based upon multiomics data sets from Streptomyces coelicolor M145, including RNA-seq, Ribo-seq, and TSS-seq, for controllable transcription and translation. A total eight sets of promoter/5′ UTR combinations, with minimal interferences of promoters on translation, were constructed using the transcription start site information, and evaluated with the GusA system. Expression of GusA could be controlled to various strengths in three different media, in a range of 0.03- to 2.4-fold, compared to that of the control, ermE*P/Shine-Dalgarno sequence. This method was applied to engineer three previously reported promo...

Systems biology for understanding and engineering of heterotrophic oleaginous microorganisms.

Abstract: Heterotrophic oleaginous microorganisms continue to draw interest as they can accumulate a large amount of lipids which is a promising feedstock for the production of biofuels and oleochemicals. Nutrient limitation, especially nitrogen limitation, is known to effectively trigger the lipid production in these microorganisms. For the aim of developing improved strains, the mechanisms behind the lipid production have been studied for a long time. Nowadays, system-level understanding of their metabolism and associated metabolic switches is attainable with modern systems biology tools. This work reviews the systems biology studies, based on (i) top-down, large-scale 'omics' tools, and (ii) bottom-up, mathematical modeling methods, on the heterotrophic oleaginous microorganisms with an emphasis on further application to metabolic engineering.

BeReTa: a systematic method for identifying target transcriptional regulators to enhance microbial production of chemicals.

Abstract: MOTIVATION Modulation of regulatory circuits governing the metabolic processes is a crucial step for developing microbial cell factories. Despite the prevalence of in silico strain design algorithms, most of them are not capable of predicting required modifications in regulatory networks. Although a few algorithms may predict relevant targets for transcriptional regulator (TR) manipulations, they have limited reliability and applicability due to their high dependency on the availability of integrated metabolic/regulatory models. RESULTS We present BeReTa (Beneficial Regulator Targeting), a new algorithm for prioritization of TR manipulation targets, which makes use of unintegrated network models. BeReTa identifies TR manipulation targets by evaluating regulatory strengths of interactions and beneficial effects of reactions, and subsequently assigning beneficial scores for the TRs. We demonstrate that BeReTa can predict both known and novel TR manipulation targets for enhanced production of various chemicals in Escherichia coli Furthermore, through a case study of antibiotics production in Streptomyces coelicolor, we successfully demonstrate its wide applicability to even less-studied organisms. To the best of our knowledge, BeReTa is the first strain design algorithm exclusively designed for predicting TR manipulation targets. AVAILABILITY AND IMPLEMENTATION MATLAB code is available at https://github.com/kms1041/BeReTa (github). CONTACT byungkim@snu.ac.krSupplementary information: Supplementary data are available at Bioinformatics online.

The role of NdgR in glycerol metabolism in Streptomyces coelicolor

Abstract: Streptomyces, which produces many pharmaceutical antibiotics and anticancer agents, is a genus of soil-dwelling bacteria with numerous regulators that control both primary and secondary metabolism. NdgR is highly conserved in Streptomyces spp. and is known to be involved in antibiotic production, tolerance against shock and physical stress, nitrogen metabolism, leucine metabolism, and N-acetylglucosamine metabolism. As another function of NdgR, we report the involvement of NdgR in glycerol metabolism in S. coelicolor. Initially, a glycerol utilization operon containing gylCABX was found to be up-regulated in an ndgR deletion mutant (BG11) grown in N-acetylglucosamine solid minimal media compared with wild-type strain (M145). BG11 produced more antibiotics with a small amount of glycerol and increased glycerol utilization, yielding higher concentrations of lactate and acetate per cell. Moreover, fatty acid production was also changed in BG11 to produce longer chain fatty acids, phenolic compounds, alkanes, and fatty alcohols. Using a gel retardation assay, NdgR was found to bind the upstream region of gylC, working as a repressor. NdgR is a second regulator of a glycerol utilization operon, for which only one regulator, GylR was previously known.