Publications by authors named "Jin Suk Lee"

82 Publications

Improving the organic solvent resistance of lipase a from Bacillus subtilis in water-ethanol solvent through rational surface engineering.

Bioresour Technol 2021 Jun 9;337:125394. Epub 2021 Jun 9.

Department of Biotechnology, The Catholic University of Korea, Bucheon-si, Gyeonggi-do 14662, Republic of Korea. Electronic address:

Given that lipase is an enzyme applicable in various industrial fields and water-miscible organic solvents are important reaction media for developing industrial-scale biocatalysis, a structure-based strategy was explored to stabilize lipase A from Bacillus subtilis in a water-ethanol cosolvent. Site-directed mutagenesis of ethanol-interacting sites resulted in 4 mutants, i.e., Ser16Gly, Ala38Gly, Ala38Thr, and Leu108Asn, which were stable in 50% ethanol and had up to 1.8-fold higher stability than the wild-type. In addition, Leu108Asn was more thermostable at 45 °C than the wild type. The results discussed in this study not only provide insights into strategies for enzyme engineering to improve organic solvent resistance but also suggest perspectives on pioneering routes for constructing enzyme-based biorefineries to produce value-added fuels and chemicals.
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http://dx.doi.org/10.1016/j.biortech.2021.125394DOI Listing
June 2021

Identification of genes involved in neuronal cell death and recovery over time in rat axotomy and neurorrhaphy models through RNA sequencing.

Mol Cell Neurosci 2021 Jun 5;113:103628. Epub 2021 May 5.

Regeneration Medicine Research Center, Yonsei University Wonju College of Medicine, Wonju, Republic of Korea. Electronic address:

Facial nerves are frequently injured during cosmetic or other types of facial surgery. However, information on the genes involved in the damage and recovery of the facial nerves is limited. Here, we aimed to identify the genes affected by facial nerve injury and repair using next-generation sequencing. We established a rat axotomy model and a parallel epineurial neurorrhaphy model, in which gene expression was analyzed from 3 days to 8 weeks after surgery. We discovered that ARRB1, SGK1, and GSK3B genes associated with neuronal cell death were upregulated in the axotomy model. In contrast, MFRP, MDK, and ACE genes involved in neural recovery and regeneration exhibited higher expression in the neurorrhaphy model. In the present study, the analysis of the big data obtained from the next-generation sequencing (RNA-seq) technology reveals that the expression of genes involved in neuronal cell death is induced during nerve damage, and those associated with neural recovery are more abundantly expressed during repair processes. These results are considered to be useful for the establishment of the treatment of related diseases and basic research in various neuroscience fields by utilizing damage and recovery mechanism of facial nerves.
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http://dx.doi.org/10.1016/j.mcn.2021.103628DOI Listing
June 2021

Recent developments and key barriers to microbial CO electrobiorefinery.

Bioresour Technol 2021 Jan 2;320(Pt A):124350. Epub 2020 Nov 2.

Gwangju Bio/Energy R&D Center, Korea Institute of Energy Research, Gwangju 61003, Republic of Korea. Electronic address:

The electrochemical conversion of CO can include renewable surplus electricity storage and CO utilisation. This review focuses on the microbial CO electrobiorefinery based on microbial electrosynthesis (MES) which merges electrochemical and microbial conversion to produce biofuels and higher-value chemicals. In this review, recent developments are discussed about bioelectrochemical conversion of CO into biofuels and chemicals in MES via microbial CO-fixation and electricity utilisation reactions. In addition, this review examines technical approaches to overcome the current limitations of MES including the following: engineering of the biocathode, application of electron mediators, and reactor optimisation, among others. An in-depth discussion of strategies for the CO electrobiorefinery is presented, including the integration of the biocathode with inorganic catalysts, screening of novel electroactive microorganisms, and metabolic engineering to improve target productivity from CO.
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http://dx.doi.org/10.1016/j.biortech.2020.124350DOI Listing
January 2021

The Role of Negative Emotions Pre- and Post-Implementation of Graphic Health Warnings: Longitudinal Evidence from South Korea.

Int J Environ Res Public Health 2020 07 27;17(15). Epub 2020 Jul 27.

Dongguk Business School, Dongguk University, Seoul 04620, Korea.

This longitudinal study investigated the factors that determine the effectiveness of graphic health warnings (GHWs) by comparing 246 South Korean smoker's responses before and after the introduction of the country's new tobacco control policy wherein GHWs were placed on all cigarette packaging. Even though introducing GHWs did not cause immediate changes in smokers' intention to quit smoking or perception of smoking's health risk, GHWs eventually motivated smokers to quit smoking when they experienced negative emotional responses to the newly introduced graphic warnings on cigarette packaging. More importantly, this study found that positive changes in smokers' perceived risk associated with smoking due to the introduction of GHWs mediated a positive relationship between changes in smokers' negative emotions (NE) from text-only warnings to graphic warnings and changes in their intention to quit smoking during the same period. Based on these results, the authors suggest that, for GHW policy to be more effective in motivating smoking cessation, the warnings need to convey images sufficiently unpleasant to induce negative emotional responses among smokers.
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http://dx.doi.org/10.3390/ijerph17155393DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7432201PMC
July 2020

Biocompatible liquid-type carbon nanodots (C-paints) as light delivery materials for cell growth and astaxanthin induction of Haematococcus pluvialis.

Mater Sci Eng C Mater Biol Appl 2020 Apr 28;109:110500. Epub 2019 Nov 28.

Division of Material Analysis and Research, Korea Basic Science Institute, Daejeon 305-333, Republic of Korea. Electronic address:

In this study, we aimed to demonstrate the feasibility of the application of biocompatible liquid type fluorescent carbon nanodots (C-paints) to microalgae by improving microalgae productivity. C-paints were prepared by a simple process of ultrasound irradiation using polyethylene glycol (PEG) as a passivation agent. The resulting C-paints exhibited a carbonyl-rich surface with good uniformity of particle size, excellent water solubility, photo-stability, fluorescence efficiency, and good biocompatibility (<10.0 mg mL of C-paints concentration). In the practical application of C-paints to microalgae culture, the most effective and optimized condition leading to growth promoting effect was observed at a C-paints concentration of 1.0 mg mL (>20% higher than the control cell content). A C-paints concentration of 1-10.0 mg mL induced an approximately >1.8 times higher astaxanthin content than the control cells. The high light delivery effect of non-cytotoxic C-paints was applied as a stress condition for H. pluvialis growth and was found to play a major role in enhancing productivity. Notably, the results from this study are an essential approach to improve astaxanthin production, which can be used in various applications because of its therapeutic effects such as cancer prevention, anti-inflammation, immune stimulation, and treatment of muscle-soreness.
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http://dx.doi.org/10.1016/j.msec.2019.110500DOI Listing
April 2020

Adipose Tissue-Derived Mesenchymal Stem Cells Suppress Growth of Huh7 Hepatocellular Carcinoma Cells via Interferon (IFN)-β-Mediated JAK/STAT1 Pathway .

Int J Med Sci 2020 18;17(5):609-619. Epub 2020 Feb 18.

Regeneration Medicine Research Center, Yonsei University Wonju College of Medicine, Wonju, Gangwon-do 26426, Republic of Korea.

Interferon (IFN)-β and/or tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) secreted by adipose tissue-derived mesenchymal stem cells (ASCs) have been proposed as key mechanistic factors in anti-cancer efficacy in lung cancer and breast cancer cells, where they act through paracrine signaling. We hypothesized that IFN-β and TRAIL produced by ASCs suppress proliferation of hepatocellular carcinoma cells (HCCs). The present study evaluated the anti-cancer effects of ASCs on HCCs . We found that indirect co-culture with ASCs diminished growth of Huh7 hepatocellular carcinoma cells with increased protein levels of p53/p21 and phosphorylated STAT1 (pSTAT1), without apoptosis. Treatment with ASC-conditioned medium (ASC-CM) also decreased growth of Huh7 cells through elevated p53/p21 and pSTAT1 signaling. ASC-CM-mediated inhibition of cell growth was neutralized in Huh7 cells treated with anti-IFN-β antibody compared to that in ASC-CM-treated Huh7 cells incubated with an anti-TRAIL antibody. Treatment with JAK1/JAK2 inhibitors recovered inhibition of growth in Huh7 cells incubated in ASC-CM or IFN-β via down-regulation of pSTAT1/p53/p21. However, treatment of IFN-β resulted in no alterations in resistance of Huh7 cells to TRAIL. Our findings suggest that ASCs decrease growth through activated STAT1-mediated p53/p21 by IFN-β, but not TRAIL, in Huh7 cells.
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http://dx.doi.org/10.7150/ijms.41354DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7085211PMC
December 2020

Elevated conversion of CO to versatile formate by a newly discovered formate dehydrogenase from Rhodobacter aestuarii.

Bioresour Technol 2020 Jun 6;305:123155. Epub 2020 Mar 6.

Gwangju Bio/Energy Research Center, Korea Institute of Energy Research (KIER), Gwangju 61003, Republic of Korea. Electronic address:

Due to climate change, recent research interests have increased towards CO utilization as a strategy to mitigate the atmospheric CO level. Herein, we aimed to explore formate dehydrogenases (FDHs) from chemoautotroph to discover an efficient and O-tolerant biocatalyst for catalyzing the CO reduction to a versatile formate. Through genome-mining and phylogenetic analysis, the FDH from Rhodobacter aestuarii (RaFDH) was newly discovered as a promising O-tolernat CO reductase and was successfully expressed in Escherichia coli. In this study, the optimum conditions and turnover rates of RaFDH were examined for CO reduction and formate oxidation. In particular, the RaFDH-driven CO reduction far surpassed the formate oxidation with a turnover rate of 48.3 and 15.6 min, respectively. The outstanding superiority of RaFDH towards CO reduction can be applicable for constructing a feasible electroenzymatic system that produce a versatile formate from CO as a cheap, abundant, and renewable resource.
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http://dx.doi.org/10.1016/j.biortech.2020.123155DOI Listing
June 2020

Pressurized cultivation strategies for improved microbial hydrogen production by Thermococcus onnurineus NA1.

Bioprocess Biosyst Eng 2020 Jun 30;43(6):1119-1122. Epub 2020 Jan 30.

Gwangju Bio/Energy R&D Center, Korea Institute of Energy Research, Gwangju, 61003, Republic of Korea.

While the hydrogen economy is receiving growing attention, research on microbial hydrogen production is also increasing. Microbial water-gas shift reaction is advantageous as it produces hydrogen from by product gas including carbon monoxide (CO). However, CO solubility in water is the bottleneck of this process by low mass transfer. Thermococcus onnurineus NA1 strain can endure a high-pressure environment and can enhance hydrogen production in a pressurized reactor by increasing CO solubility. As CO causes cell toxicity, two important factors, pressure and input gas flow rate, should be considered for process control during cultivation. Hence, we employed different operational strategies for enhancing hydrogen production and obtained 577 mmol/L/h of hydrogen productivity. This is the highest hydrogen productivity reported to date from microbial water-gas shift reaction.
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http://dx.doi.org/10.1007/s00449-020-02291-yDOI Listing
June 2020

Rapid induction of edible lipids in Chlorella by mild electric stimulation.

Bioresour Technol 2019 Nov 2;292:121950. Epub 2019 Aug 2.

School of Chemical & Biomolecular Engineering, Pusan National University, Busan 46241, Republic of Korea. Electronic address:

In this work, a new stress-based method for rapid induction of triacylglycerol (TAG) and total and polyunsaturated fatty acid accumulations in Chlorella sp. by mild electric stimulation is presented. When a cathodic current of 31 mA (voltage: 4 V) was applied to the algal cells for 4 h, the TAG content of the electro-treated cells was sharply increased to a level 2.1 times that of the untreated control. The contents of the polyunsaturated linoleic (C18:2n6) and linolenic (C18:3n3) acids in the electro-treated cells were also 36 and 57% higher than those in the untreated cells, respectively. Cyclic voltammetry and various biochemical analyses indicate that TAG and fatty acid formations are electro-stimulated via de novo fatty acid biosynthesis and metabolic transformation in the Chlorella cells.
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http://dx.doi.org/10.1016/j.biortech.2019.121950DOI Listing
November 2019

A perspective on the biotechnological applications of the versatile tyrosinase.

Bioresour Technol 2019 Oct 29;289:121730. Epub 2019 Jun 29.

Bio/Energy R&D Center, Korea Institute of Energy Research (KIER), Gwangju 61003, Republic of Korea. Electronic address:

Tyrosinase (E.C. 1.14.18. 1) is a type of Cu-containing oxidoreductase which has bifunctional activity for various phenolic substrates: ortho-hydroxylation of monophenols to diphenols (a cresolase activity) and oxidation of diphenols to quinones (a catecholase activity). Based on the broad substrate spectrum, tyrosinase has been used in bioremediation of phenolic pollutants, constructing biosensors for identifying phenolic compounds, and L-DOPA synthesis. Furthermore, not only tyrosinase has been used to produce useful polyphenol derivatives, but also it is recently revealed that the promiscuous activity of tyrosinase is closely related with delignification in the biorefinery. Accordingly, tyrosinase might be a potential biocatalyst for industrial applications (e.g., electroenzymatic L-DOPA production, but its long-term stability and reusability should be further explored. In this review, we emphasize the versatility of tyrosinase, which includes conventional applications, and suggest new perspectives as an industrial biocatalyst (e.g., electroenzymatic L-DOPA production). Especially, this review focuses on and comprehensively discusses recent innovative studies.
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http://dx.doi.org/10.1016/j.biortech.2019.121730DOI Listing
October 2019

MKP1 acts as a key modulator of stomatal development.

Plant Signal Behav 2019 13;14(7):1604017. Epub 2019 Apr 13.

a Department of Biology , Concordia University , Montreal , Quebec , Canada.

The MAPK signaling cascade is universal among eukaryotes and mediates a variety of environmental and developmental responses. Two Arabidopsis MAPKs, MPK3 and MPK6, have been shown to be activated by various stimuli and suggested as a convergence point of different signaling pathways. It is known that these MAPKs, MPK3/MPK6, control the discrete stages of stomatal development in Arabidopsis, but how they are regulated and how the same MAPK components can achieve signaling specificity is largely unknown. We recently demonstrated that MAP Kinase Phosphatase 1 (MKP1) promotes stomatal differentiation by suppressing activation of MPK3/MPK6 in the stomatal lineage. By expressing MKP1 in discrete stomatal precursor cell types, we further identified that MKP1 plays an important role at the early stage of stomatal development for the cell fate transition leading to stomatal differentiation. While MKP1 was previously known as a key regulator of environmental stress responses, our data illustrate a novel role of MKP1 in plant development: it acts as one of the specificity-determining regulators of MAPK signaling to enforce proper stomatal development in Arabidopsis.
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http://dx.doi.org/10.1080/15592324.2019.1604017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6619980PMC
May 2020

Critical Point Drying: An Effective Drying Method for Direct Measurement of the Surface Area of a Pretreated Cellulosic Biomass.

Polymers (Basel) 2018 Jun 17;10(6). Epub 2018 Jun 17.

Gwangju Bioenergy R&D Center, Korea Institute of Energy Research, 25 Samso-ro270beongil, Buk-gu, Gwangju 61003, Korea.

The surface area and pore size distribution of samples that were pretreated by different methods were determined by the Brunauer⁻Emmett⁻Teller (BET) technique. Three methods were applied to prepare cellulosic biomass samples for the BET measurements, air, freeze, and critical point drying (CPD). The air and freeze drying caused a severe collapse of the biomass pore structures, but the CPD effectively preserved the biomass morphology. The surface area of the CPD prepared samples were determined to be 58⁻161 m²/g, whereas the air and freeze dried samples were 0.5⁻1.3 and 1.0⁻2.4 m²/g, respectively. The average pore diameter of the CPD prepared samples were 61⁻70 Å. The CPD preserved the sample morphology by replacing water with a non-polar solvent, CO₂ fluid, which prevented hydrogen bond reformation in the cellulose.
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http://dx.doi.org/10.3390/polym10060676DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6404156PMC
June 2018

Anti-EGF Receptor Aptamer-Guided Co-Delivery of Anti-Cancer siRNAs and Quantum Dots for Theranostics of Triple-Negative Breast Cancer.

Theranostics 2019 25;9(3):837-852. Epub 2019 Jan 25.

Department of Biomedical Laboratory Science, Yonsei University, Wonju, Republic of Korea.

Many aptamers have been evaluated for their ability as drug delivery vehicles to target ligands, and a variety of small interfering RNAs (siRNAs) have been tested for their anti-cancer properties. However, since these two types of molecules have similar physicochemical properties, it has so far been difficult to formulate siRNA-encapsulating carriers guided by aptamers. Here, we propose aptamer-coupled lipid nanocarriers encapsulating quantum dots (QDs) and siRNAs for theragnosis of triple-negative breast cancer (TNBC). Hydrophobic QDs were effectively incorporated into lipid bilayers, and then therapeutic siRNAs were complexed with QD-lipid nanocarriers (QLs). Finally, anti-EGFR aptamer-lipid conjugates were inserted into the QLs for TNBC targeting (aptamo-QLs). TNBC-targeting aptamo-QLs were directly compared to anti-EGFR antibody-coupled immuno-QLs. The delivery of therapeutic siRNAs and QDs to target cells was assessed by flow cytometry and confocal microscopy. The targeting of siRNAs to tumors and their therapeutic efficacy were evaluated in mice carrying MDA-MB-231 tumors. Both types of EGFR-targeting QLs showed enhanced delivery to target cancer cells, resulting in more effective gene silencing and enhanced tumor imaging compared to non-targeting control QLs. Moreover, combinatorial therapy with Bcl-2 and PKC-ι siRNAs loaded into the anti-EGFR QLs was remarkably effective in inhibiting tumor growth and metastasis. In general, the aptamo-QLs showed competitive delivery and therapeutic efficacy compared to immuno-QLs under the same experimental conditions. Our results show that the anti-EGFR aptamer-guided lipid carriers may be a potential theranostic delivery vehicle for RNA interference and fluorescence imaging of TNBCs.
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http://dx.doi.org/10.7150/thno.30228DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6376474PMC
December 2019

High-efficiency cell disruption and astaxanthin recovery from Haematococcus pluvialis cyst cells using room-temperature imidazolium-based ionic liquid/water mixtures.

Bioresour Technol 2019 Feb 23;274:120-126. Epub 2018 Nov 23.

Climate Change Research Division, Korea Institute of Energy Research, Daejeon 34129, Republic of Korea. Electronic address:

Energy-saving, high-efficiency cell disruption is a critical step for recovery of thermolabile antioxidant astaxanthin from Haematococcus pluvialis cyst cells of rigid cell-wall structure. In this study, as room-temperature green solvents, 10 types of 1-ethyl-3-methylimidazolium ([Emim])-based ionic liquids (ILs) were compared and evaluated for their abilities to disrupt H. pluvialis cyst cells for astaxanthin/lipid extraction. Among the 10 ILs tested, 3 [Emim]-based ILs with HSO, CHSO, and (CFSO)N anions were selected based on astaxanthin/lipid extraction performance and synthesis cost. When pretreated with IL/water mixtures, intact cyst cells were significantly torn, broken or shown to release cytoplasmic components, thereby facilitating subsequent separation of astaxanthin/lipid by hexane. However, excess IL pretreatments at high temperature/IL dosages and longer incubation times significantly deteriorated lipid and/or astaxanthin. Under optimized mild conditions (6.7% (v/v) IL in water solution, 30 °C, 60 min), almost complete astaxanthin recoveries (>99%) along with moderate lipid extractions (∼82%) could be obtained.
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http://dx.doi.org/10.1016/j.biortech.2018.11.082DOI Listing
February 2019

MAP KINASE PHOSPHATASE1 Controls Cell Fate Transition during Stomatal Development.

Plant Physiol 2018 09 12;178(1):247-257. Epub 2018 Jul 12.

Department of Biology, Concordia University, Montreal, Quebec H4B 1R6, Canada

Stomata on the plant epidermis control gas and water exchange and are formed by MAPK-dependent processes. Although the contribution of MAP KINASE3 (MPK3) and MPK6 (MPK3/MPK6) to the control of stomatal patterning and differentiation in Arabidopsis () has been examined extensively, how they are inactivated and regulate distinct stages of stomatal development is unknown. Here, we identify a dual-specificity phosphatase, MAP KINASE PHOSPHATASE1 (MKP1), which promotes stomatal cell fate transition by controlling MAPK activation at the early stage of stomatal development. Loss of function of creates clusters of small cells that fail to differentiate into stomata, resulting in the formation of patches of pavement cells. We show that MKP1 acts downstream of YODA (a MAPK kinase kinase) but upstream of MPK3/MPK6 in the stomatal signaling pathway and that MKP1 deficiency causes stomatal signal-induced MAPK hyperactivation in vivo. By expressing MKP1 in the three discrete cell types of stomatal lineage, we further identified that MKP1-mediated deactivation of MAPKs in early stomatal precursor cells directs cell fate transition leading to stomatal differentiation. Together, our data reveal the important role of MKP1 in controlling MAPK signaling specificity and cell fate decision during stomatal development.
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http://dx.doi.org/10.1104/pp.18.00475DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6130035PMC
September 2018

Antagonistic interactions between two MAP kinase cascades in plant development and immune signaling.

EMBO Rep 2018 07 22;19(7). Epub 2018 May 22.

Department of Botany, University of British Columbia, Vancouver, BC, Canada

Mitogen-activated protein kinase (MAPK) signaling plays important roles in diverse biological processes. In , MPK3/MPK6, MKK4/MKK5, and the MAPKKK YODA (YDA) form a MAPK pathway that negatively regulates stomatal development. Brassinosteroid (BR) stimulates this pathway to inhibit stomata production. In addition, MPK3/MPK6 and MKK4/MKK5 also serve as critical signaling components in plant immunity. Here, we report that MAPKKK3/MAPKKK5 form a kinase cascade with MKK4/MKK5 and MPK3/MPK6 to transduce defense signals downstream of multiple plant receptor kinases. Loss of MAPKKK3/MAPKKK5 leads to reduced activation of MPK3/MPK6 in response to different pathogen-associated molecular patterns (PAMPs) and increased susceptibility to pathogens. Surprisingly, developmental defects caused by silencing of are suppressed in the double mutant. On the other hand, loss of YDA or blocking BR signaling leads to increased PAMP-induced activation of MPK3/MPK6. These results reveal antagonistic interactions between a developmental MAPK pathway and an immune signaling MAPK pathway.
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http://dx.doi.org/10.15252/embr.201745324DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6030703PMC
July 2018

Recent developments and key barriers to advanced biofuels: A short review.

Bioresour Technol 2018 Jun 21;257:320-333. Epub 2018 Feb 21.

Gwangju Bioenergy R&D Center, Korea Institute of Energy Research, Gwangju 61003, Republic of Korea. Electronic address:

Biofuels are regarded as one of the most viable options for reduction of CO emissions in the transport sector. However, conventional plant-based biofuels (e.g., biodiesel, bioethanol)'s share of total transportation-fuel consumption in 2016 was very low, about 4%, due to several major limitations including shortage of raw materials, low CO mitigation effect, blending wall, and poor cost competitiveness. Advanced biofuels such as drop-in, microalgal, and electro biofuels, especially from inedible biomass, are considered to be a promising solution to the problem of how to cope with the growing biofuel demand. In this paper, recent developments in oxy-free hydrocarbon conversion via catalytic deoxygenation reactions, the selection of and lipid-content enhancement of oleaginous microalgae, electrochemical biofuel conversion, and the diversification of valuable products from biomass and intermediates are reviewed. The challenges and prospects for future development of eco-friendly and economically advanced biofuel production processes also are outlined herein.
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http://dx.doi.org/10.1016/j.biortech.2018.02.089DOI Listing
June 2018

A comparison of various lignin-extraction methods to enhance the accessibility and ease of enzymatic hydrolysis of the cellulosic component of steam-pretreated poplar.

Biotechnol Biofuels 2017 19;10:157. Epub 2017 Jun 19.

Forest Products Biotechnology/Bioenergy Group, Department of Wood Science, Faculty of Forestry, University of British Columbia, 2424 Main Mall, Vancouver, BC V6T 1Z4 Canada.

Background: Current single-stage delignification-pretreatment technologies to overcome lignocellulosic biomass recalcitrance are usually achieved at the expense of compromising the recovery of the polysaccharide components, particularly the hemicellulose fraction. One way to enhance overall sugar recovery is to tailor an efficient two-stage pretreatment that can pre-extract the more labile hemicellulose component before subjecting the cellulose-rich residual material to a second-stage delignification process. Previous work had shown that a mild steam pretreatment could recover >65% of the hemicellulose from poplar while limiting the acid-catalysed condensation of lignin. This potentially allowed for subsequent lignin extraction using various lignin solvents to produce a more accessible cellulosic substrate.

Results: A two-stage approach using steam and/or solvent pretreatment was assessed for its ability to separate hemicellulose and lignin from poplar wood chips while providing a cellulose-rich fraction that could be readily hydrolysed by cellulase enzymes. An initial steam-pretreatment stage was performed over a range of temperatures (160-200 °C) using an equivalent severity factor of 3.6. A higher steam temperature of 190 °C applied over a shorter residence time of 10 min effectively solubilized and recovered 75% of the hemicellulose while enhancing the ability of various solvents [deep eutectic solvent (DES), ethanol organosolv, soda/anthraquinone (soda/AQ) or a hydrotrope] to extract lignin in a second stage. When the second-stage treatments were compared, the mild DES treatment (lactic acid and betaine) at 130 °C, removed comparable amounts of lignin with higher selectivity than did the soda/AQ and organosolv pretreatments at 170 °C. However, the cellulose-rich substrates obtained after the second-stage organosolv and soda/AQ pretreatments showed the highest cellulose accessibility, as measured by the Simon's staining technique. They were also the most susceptible to subsequent enzymatic hydrolysis.

Conclusions: The second-stage pretreatments varied in their ability to solubilize and extract the lignin component of steam-pretreated poplar while enhancing the enzymatic hydrolysis of the resulting cellulose-rich residual fractions. Although DES extraction was more selective in extracting lignin from the steam-pretreated substrates, the organosolv and soda/AQ post treatments disrupted the cellulose structure to a greater extent while enhancing the ease of enzymatic hydrolysis. Graphical abstractEffective hemicellulose removal via steam pretreatment followed by subsequent lignin extraction under acidic, alkaline or solvolytic conditions results in a highly accessible, more readily hydrolysed cellulose fraction.
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http://dx.doi.org/10.1186/s13068-017-0846-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5477284PMC
June 2017

Purification of Plant Receptor Kinases from Plant Plasma Membranes.

Authors:
Jin Suk Lee

Methods Mol Biol 2017 ;1621:47-56

Department of Biology, Concordia University, 7141 Sherbrooke W., Montreal, QC, Canada, H4B 1R6.

Receptor kinases play a central role in various biological processes, but due to their low abundance and highly hydrophobic and dynamic nature, only a few of them have been functionally characterized, and their partners and ligands remain unidentified. Receptor protein extraction and purification from plant tissues is one of the most challenging steps for the success of various biochemical analyses to characterize their function. Immunoprecipitation is a widely used and selective method for enriching or purifying a specific protein. Here we describe two different optimized protein purification protocols, batch and on-chip immunoprecipitation, which efficiently isolate plant membrane receptor kinases for functional analysis.
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http://dx.doi.org/10.1007/978-1-4939-7063-6_5DOI Listing
March 2018

Peritoneal dialysis-related peritonitis due to Halomonas hamiltonii: A first case report.

Medicine (Baltimore) 2016 Nov;95(47):e5424

Department of Internal Medicine, Dongguk University Lee's Renal Clinic, Gyeongju Korean Collection for Type Cultures, Biological Resource Center, Korea Research Institute of Bioscience and Biotechnology, Jeongeup Department of Laboratory Medicine, Dongguk University, Gyeongju Hogye 365 Medical Clinic, Ulsan, Korea.

Introduction: Halomonas hamiltonii is a Gram-negative, halophilic, motile, and nonspore-forming rod bacterium. Although most Halomonas sp. are commonly found in saline environments, it has rarely been implicated as a cause of human infection. Herein, the authors present a case report of continuous ambulatory peritoneal dialysis (CAPD)-related peritonitis attributed to H hamiltonii.

Case Presentation: An 82-year-old male patient who had been receiving CAPD therapy presented to an emergency department with complaints of abdominal pain and cloudy dialysate that had persisted for 2 days. The peritoneal dialysate was compatible with CAPD peritonitis, with white blood cell count of peritoneal effluent of 810/mm and neutrophils predominated (60%). Two days after culture on blood agar medium, nonhemolytic pink mucoid colonies showed, with cells showing Gram-negative, nonspore-forming rods with a few longer and larger bacilli than usual were found. We also performed biochemical tests and found negative responses in K/K on the triple sugar iron test and H2S and equivocal (very weak) response in the motility test, but positive responses to catalase, oxidase, and urease tests. The partial sequence of the 16S rRNA gene of a bacterium detected by peritoneal fluid culture was utilized for a Basic Local Alignment Search Tool search, which revealed that the organism was H hamiltonii. Intraperitoneal antibiotics were administered for 21 days, and the patient was discharged without clinical problems.

Conclusion: We present here the first case report of CAPD-related peritonitis caused by H hamiltonii, which was identified using molecular biological techniques. Although guidelines do not exist for the treatment of infections caused by this organism, conventional treatment for Gram-negative organisms could be effective.
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http://dx.doi.org/10.1097/MD.0000000000005424DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5134875PMC
November 2016

Fine-Tuning Development Through Antagonistic Peptides: An Emerging Theme.

Trends Plant Sci 2016 12 18;21(12):991-993. Epub 2016 Oct 18.

Department of Plant Systems Biology, VIB, B-9052 Ghent, Belgium; Department of Plant Biotechnology and Bioinformatics, Ghent University, B-9052 Ghent, Belgium. Electronic address:

Peptide ligand-receptor kinase interactions have emerged as a key component of plant growth and development. Now, highly related small signaling peptides have been shown to act antagonistically on the same receptor kinase, providing new insights into how plants optimize developmental processes using competitive peptides.
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http://dx.doi.org/10.1016/j.tplants.2016.10.003DOI Listing
December 2016

A Secreted Peptide and Its Receptors Shape the Auxin Response Pattern and Leaf Margin Morphogenesis.

Curr Biol 2016 09 1;26(18):2478-2485. Epub 2016 Sep 1.

Institute of Transformative Bio-Molecules (WPI-ITbM), Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan; Division of Biological Science, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8602, Japan. Electronic address:

Secreted peptides mediate intercellular communication [1, 2]. Several secreted peptides in the EPIDERMAL PATTERNING FACTOR-LIKE (EPFL) family regulate morphogenesis of tissues, such as stomata and inflorescences in plants [3-15]. The biological functions of other EPFL family members remain unknown. Here, we show that the EPFL2 gene is required for growth of leaf teeth. EPFL2 peptide physically interacts with ERECTA (ER) family receptor-kinases and, accordingly, the attenuation of ER family activities leads to formation of toothless leaves. During the tooth growth process, responses to the phytohormone auxin are maintained at tips of the teeth to promote their growth [16-19]. In the growing tooth tip of epfl2 and multiple er family mutants, the auxin response becomes broader. Conversely, overexpression of EPFL2 diminishes the auxin response, indicating that the EPFL2 signal restricts the auxin response to the tooth tip. Interestingly, the tip-specific auxin response in turn organizes characteristic expression patterns of ER family and EPFL2 by enhancing ER family expression at the tip while eliminating the EPFL2 expression from the tip. Our findings identify the novel ligand-receptor pairs promoting the tooth growth, and further reveal a feedback circuit between the peptide-receptor system and auxin response as a mechanism for maintaining proper auxin maxima during leaf margin morphogenesis.
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http://dx.doi.org/10.1016/j.cub.2016.07.014DOI Listing
September 2016

Pretreatment of biomass.

Bioresour Technol 2016 Jan;199

Biotechnology Division, CSIR-National Institute for Interdisciplinary Science and Technology, Trivandrum, India. Electronic address:

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http://dx.doi.org/10.1016/j.biortech.2015.10.092DOI Listing
January 2016

The influence of lignin on steam pretreatment and mechanical pulping of poplar to achieve high sugar recovery and ease of enzymatic hydrolysis.

Bioresour Technol 2016 Jan 11;199:135-141. Epub 2015 Sep 11.

Forest Products Biotechnology/Bioenergy Group, University of British Columbia, Faculty of Forestry, British Columbia, Canada.

With the goal of enhancing overall carbohydrate recovery and reducing enzyme loading refiner mechanical pulping and steam pretreatment (210°C, 5 min) were used to pretreat poplar wood chips. Neutral sulphonation post-treatment indicated that, although the lignin present in the steam pretreated substrate was less reactive, the cellulose-rich, water insoluble component was more accessible to cellulases and Simons stain. This was likely due to lignin relocation as the relative surface lignin measured by X-ray photoelectron spectroscopy increased from 0.4 to 0.8. The integration of sulphite directly into steam pretreatment resulted in the solubilisation of 60% of the lignin while more than 80% of the carbohydrate present in the original substrate was recovered in the water insoluble fraction after Na2CO3 addition. More than 80% of the sugars present in the original cellulose and xylan could be recovered after 48 h using an enzyme loading of 20 mg protein/g cellulose at a 10% substrate concentration.
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http://dx.doi.org/10.1016/j.biortech.2015.09.019DOI Listing
January 2016

Cell-wall disruption and lipid/astaxanthin extraction from microalgae: Chlorella and Haematococcus.

Bioresour Technol 2016 Jan 31;199:300-310. Epub 2015 Aug 31.

Biomass and Waste Energy Laboratory, Korea Institute of Energy Research, 152 Gajeong-ro, Yuseong-gu, Daejeon 305-343, Republic of Korea. Electronic address:

Recently, biofuels and nutraceuticals produced from microalgae have emerged as major interests, resulting in intensive research of the microalgal biorefinery process. In this paper, recent developments in cell-wall disruption and extraction methods are reviewed, focusing on lipid and astaxanthin production from the biotechnologically important microalgae Chlorella and Haematococcus, respectively. As a common, critical bottleneck for recovery of intracellular components such as lipid and astaxanthin from these microalgae, the composition and structure of rigid, thick cell-walls were analyzed. Various chemical, physical, physico-chemical, and biological methods applied for cell-wall breakage and lipid/astaxanthin extraction from Chlorella and Haematococcus are discussed in detail and compared based on efficiency, energy consumption, type and dosage of solvent, biomass concentration and status (wet/dried), toxicity, scalability, and synergistic combinations. This report could serve as a useful guide to the implementation of practical downstream processes for recovery of valuable products from microalgae including Chlorella and Haematococcus.
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http://dx.doi.org/10.1016/j.biortech.2015.08.107DOI Listing
January 2016

Molecular Framework of a Regulatory Circuit Initiating Two-Dimensional Spatial Patterning of Stomatal Lineage.

PLoS Genet 2015 Jul 23;11(7):e1005374. Epub 2015 Jul 23.

Howard Hughes Medical Institute, University of Washington, Seattle, Washington, United States of America; Department of Biology, University of Washington, Seattle, Washington, United States of America.

Stomata, valves on the plant epidermis, are critical for plant growth and survival, and the presence of stomata impacts the global water and carbon cycle. Although transcription factors and cell-cell signaling components regulating stomatal development have been identified, it remains unclear as to how their regulatory interactions are translated into two-dimensional patterns of stomatal initial cells. Using molecular genetics, imaging, and mathematical simulation, we report a regulatory circuit that initiates the stomatal cell-lineage. The circuit includes a positive feedback loop constituting self-activation of SCREAMs that requires SPEECHLESS. This transcription factor module directly binds to the promoters and activates a secreted signal, EPIDERMAL PATTERNING FACTOR2, and the receptor modifier TOO MANY MOUTHS, while the receptor ERECTA lies outside of this module. This in turn inhibits SPCH, and hence SCRMs, thus constituting a negative feedback loop. Our mathematical model accurately predicts all known stomatal phenotypes with the inclusion of two additional components to the circuit: an EPF2-independent negative-feedback loop and a signal that lies outside of the SPCH•SCRM module. Our work reveals the intricate molecular framework governing self-organizing two-dimensional patterning in the plant epidermis.
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http://dx.doi.org/10.1371/journal.pgen.1005374DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4512730PMC
July 2015

Co-Immunoprecipitation of Membrane-Bound Receptors.

Arabidopsis Book 2015 3;13:e0180. Epub 2015 Jun 3.

Howard Hughes Medical Institute, University of Washington, Seattle, WA 98195 ; Department of Biology, University of Washington, Seattle, WA 98195.

The study of cell-surface receptor dynamics is critical for understanding how cells sense and respond to changing environments. Therefore, elucidating the mechanisms by which signals are perceived and communicated into the cell is necessary to understand immunity, development, and stress. Challenges in testing interactions of membrane-bound proteins include their dynamic nature, their abundance, and the complex dual environment (lipid/soluble) in which they reside. Co-Immunoprecipitation (Co-IP) of tagged membrane proteins is a widely used approach to test protein-protein interaction in vivo. In this protocol we present a method to perform Co-IP using enriched membrane proteins in isolated microsomal fractions. The different variations of this protocol are highlighted, including recommendations and troubleshooting guides in order to optimize its application. This Co-IP protocol has been developed to test the interaction of receptor-like kinases, their interacting partners, and peptide ligands in stable Arabidopsis thaliana lines, but can be modified to test interactions in transiently expressed proteins in tobacco, and potentially in other plant models, or scaled for large-scale protein-protein interactions at the membrane.
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http://dx.doi.org/10.1199/tab.0180DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4470539PMC
June 2015

Competitive binding of antagonistic peptides fine-tunes stomatal patterning.

Nature 2015 Jun 17;522(7557):439-43. Epub 2015 Jun 17.

1] Howard Hughes Medical Institute, University of Washington, Seattle, Washington 98195, USA [2] Department of Biology, University of Washington, Seattle, Washington 98195, USA.

During development, cells interpret complex and often conflicting signals to make optimal decisions. Plant stomata, the cellular interface between a plant and the atmosphere, develop according to positional cues, which include a family of secreted peptides called epidermal patterning factors (EPFs). How these signalling peptides orchestrate pattern formation at a molecular level remains unclear. Here we report in Arabidopsis that Stomagen (also called EPF-LIKE9) peptide, which promotes stomatal development, requires ERECTA (ER)-family receptor kinases and interferes with the inhibition of stomatal development by the EPIDERMAL PATTERNING FACTOR 2 (EPF2)-ER module. Both EPF2 and Stomagen directly bind to ER and its co-receptor TOO MANY MOUTHS. Stomagen peptide competitively replaced EPF2 binding to ER. Furthermore, application of EPF2, but not Stomagen, elicited rapid phosphorylation of downstream signalling components in vivo. Our findings demonstrate how a plant receptor agonist and antagonist define inhibitory and inductive cues to fine-tune tissue patterning on the plant epidermis.
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http://dx.doi.org/10.1038/nature14561DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4532310PMC
June 2015

Hydrothermal acid treatment for sugar extraction from Golenkinia sp.

Bioresour Technol 2015 Aug 6;190:408-11. Epub 2015 May 6.

Biomass and Waste Energy Laboratory, Korea Institute of Energy Research, 152 Gajeong-ro, Yuseong-gu, Daejeon 305-343, Republic of Korea. Electronic address:

In this study, hydrothermal acid treatment for efficient recovery of sugar from Golenkinia sp. was investigated. The initial glucose and XMG (xylose, mannose, and galactose) contents of a prepared Golenkinia sp. solution (40g/L) were 15.05 and 5.24g/L, respectively. The microalgal cell walls were hydrolyzed, for sugar recovery, by enzymatic saccharification and/or hydrothermal acid treatment. Among the various hydrothermal acid treatment conditions, the most optimal were the 2.0% H2SO4 concentration at 150°C for 15min, under which the glucose- and XMG-extraction yields were 71.7% and 64.9%, respectively. By pH 4.8, 50°C enzymatic hydrolysis after optimal hydrothermal acid treatment, the glucose- and XMG-extraction yields were additionally increased by 8.3% and 0.8%, respectively. After hydrothermal acid treatment, the combination with the enzymatic hydrolysis process improved the total sugar yield of Golenkinia sp. to 75.4%.
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http://dx.doi.org/10.1016/j.biortech.2015.04.121DOI Listing
August 2015

Types of subtalar joint facets.

Surg Radiol Anat 2015 Aug 31;37(6):629-38. Epub 2015 Mar 31.

Department of Anatomy, Yonsei University Wonju College of Medicine, 20 Ilsan-ro, Wonju, Kangwon-Do, Republic of Korea.

Articular facets of the clinical subtalar joint (CSTJ) were analyzed using a total of 118 (right 57, left 61) dry, paired calcanei and tali from 68 Korean adult cadavers. The CSTJ facets were classified into the following three types depending on their continuity: type A, all three facets are separated; type B, the anterior and middle facets are partially connected; and type C, the anterior and middle facets are fused to form a single facet. The continuity between the anterior and middle facets was represented by the degree of separation (DS), which ranged between 2.00 (type A) and 1.00 (type C). Type A was most common (39.0 %) in calcanei and rarest (11.0 %) in tali. Matching of calcaneus-talus pairs yielded five combined types: A-A (11.0 %), A-B (28.0 %), B-B (18.6 %), B-C (13.6 %), and C-C (28.8 %). The mean DS was slightly greater in calcanei (1.53) than in tali (1.32), and decreased in the order of types A-A, A-B, B-B, B-C, and C-C. The intersecting angles between the anterior and middle facets, which are related to the mobility of the CSTJ, were inversely related to the DS. These findings indicate that the anterior and middle facets are fused more frequently in tali than in calcanei, and combinations of different CSTJ facet types (A-B, B-C) exist over 40 % of feet. Our results indicate that types with a smaller DS (such as B-C and C-C) are relatively mobile but less stable compared to those with a greater DS (such as A-A and A-B).
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http://dx.doi.org/10.1007/s00276-015-1472-1DOI Listing
August 2015