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Chanyong Lee

Bio: Chanyong Lee is an academic researcher from Michigan State University. The author has contributed to research in topics: Isomerase & Glucose-6-phosphate isomerase. The author has an hindex of 7, co-authored 11 publications receiving 429 citations.

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Journal ArticleDOI
TL;DR: Two thermophilic, anaerobic, xylan-degrading bacteria, strains B6A-RIT (T = type strain) and LX-11T, were isolated from Frying Pan Springs in Yellowstone National Park as discussed by the authors.
Abstract: Two thermophilic, anaerobic, xylan-degrading bacteria, strains B6A-RIT (T = type strain) and LX-11T, were isolated from Frying Pan Springs in Yellowstone National Park. These organisms grew chemoorganotrophically by utilizing xylan and starch but not cellulose, as well as a number of di- and monosaccharides, including glucose and xylose. Both organisms had the same optimum temperature and pH for growth (60°C and pH 6.0). The fermentation products included acetate, ethanol, lactate, CO2, and H2. Both organisms were rod shaped and deposited sulfur on their cells. The major difference between the two isolates was in spore formation; strain LX-11T sporulated, whereas strain B6A-RIT were compared with other thermophilic, anaerobic, xylan-degrading bacteria by performing DNA-DNA hybridizations and total protein analyses in order to determine the relationships of these organisms. Three different groups were identified, and new taxonomic assignments are proposed. Clostridium thermocellum LQRI was least closely related to the other seven strains studied and is placed in group I, retaining its original taxonomic assignment. Clostridium thermosulfurogenes 4BT and new isolates B6A-RIT are closely related and fall into group II, for which the new genus Thermoanaerobacterium is proposed. Isolate LX-11T is designated Thermoanaerobacterium xylanolyticum sp. nov., and isolate B6A-RIT is designated Thermoanaerobacterium saccharolyticum sp. nov. Thermoanaerobacterium thermosulfurigenes 4BT (originally C. thermosulfurogenes) is the type strain of the type species of the genus. Group III strains are placed in the genus Thermoanaerobacter; this group includes Thermoanaerobacter ethanolicus JW200T, Clostridium thermohydrosulfuricum 39E and E100-69T, and Thermoanaerobium brockii HTD4T. Thermoanaerobium brockii HTD4T is renamed Thermoanaerobacter thermohydrosulfuricus comb. nov. C. thermohydrosulfuricum 39E is nearly identical to Thermoanaerobacter ethanolicus JW200T, and these organisms are considered members of the same species. Therefore, C. thermohydrosulfuricum 39E is renamed Thermoanaerobacter ethanolicus 39E; strain JW200 is the type strain of Thermoanaerobacter ethanolicus.

216 citations

Journal ArticleDOI
TL;DR: Results suggest that the enzymatic sugar isomerization does not involve a histidine-catalyzed proton transfer mechanism and, rather, essential histidine functions to stabilize the transition state by hydrogen bonding to the C5 hydroxyl group of the substrate and this enables a metal-Catalyzed hydride shift from C2 to C1.

86 citations

Journal ArticleDOI
TL;DR: A thermoanaerobe grown in TYE-starch medium at 60 degrees C produced both extra- and intracellular pullulanase and amylase activities, which did not show any metal ion activity, and both activities were inhibited by beta- and gamma- cyclodextrins but not by alpha-cyclodextrin.
Abstract: A thermoanaerobe (Thermoanaerobacter sp.) grown in TYE-starch (0.5%) medium at 60°C produced both extra- and intracellular pullulanase (1.90 U/ml) and amylase (1.19 U/ml) activities. Both activities were produced at high levels on a variety of carbon sources. The temperature and pH optima for both pullulanase and amylase activities were 75°C and pH 5.0, respectively. Both the enzyme activities were stable up to 70°C (without substrate) and at pH 4.5 to 5.0. The half-lives of both enzyme activities were 5 h at 70°C and 45 min at 75°C. The enzyme activities did not show any metal ion activity, and both activities were inhibited by β- and γ-cyclodextrins but not by α-cyclodextrin. A single amylolytic pullulanase responsible for both activities was purified to homogeneity by DEAE-Sepharose CL-6B column chromatography, gel filtration using high-pressure liquid chromatography, and pullulan-Sepharose affinity chromatography. It was a 450,000-molecular-weight glycoprotein composed of two equivalent subunits. The pullulanase cleaved pullulan in α1,6 linkages and produced multiple saccharides from cleavage of α-1,4 linkages in starch. The Kms for pullulan and soluble starch were 0.43 and 0.37 mg/ml, respectively.

48 citations

Journal ArticleDOI
TL;DR: A new assay method for thermostable glucose isomerase activity on agar plates, using a top agar mixture containing fructose, glucose oxidase, peroxidase, and benzidine, was developed and displayed properties identical to those of the native enzyme purified from C. thermosulfurogenes.
Abstract: The gene that encodes thermostable glucose isomerase in Clostridium thermosulfurogenes was cloned by complementation of glucose isomerase activity in a xylA mutant of Escherichia coli. A new assay method for thermostable glucose isomerase activity on agar plates, using a top agar mixture containing fructose, glucose oxidase, peroxidase, and benzidine, was developed. One positive clone, carrying plasmid pCGI38, was isolated from a cosmid library of C. thermosulfurogenes DNA. The plasmid was further subcloned into a Bacillus cloning vector, pTB523, to generate shuttle plasmid pMLG1, which is able to replicate in both E. coli and Bacillus subtilis. Expression of the thermostable glucose isomerase gene in both species was constitutive, whereas synthesis of the enzyme in C. thermosulfurogenes was inducible by D-xylose. B. subtilis and E. coli produced higher levels of thermostable glucose isomerase (1.54 and 0.46 U/mg of protein, respectively) than did C. thermosulfurogenes (0.29 U/mg of protein). The glucose isomerases synthesized in E. coli and B. subtilis were purified to homogeneity and displayed properties (subunit Mr, 50,000; tetrameric molecular structure; thermostability; metal ion requirement; and apparent temperature and pH optima) identical to those of the native enzyme purified from C. thermosulfurogenes. Simple heat treatment of crude extracts from E. coli and B. subtilis cells carrying the recombinant plasmid at 85 degrees C for 15 min generated 80% pure glucose isomerase. The maximum conversion yield of glucose (35%, wt/wt) to fructose with the thermostable glucose isomerase (10.8 U/g of dry substrate) was 52% at pH 7.0 and 70 degrees C.

35 citations


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TL;DR: This review concentrates on the remarkable thermostability of hyperthermophilic enzymes, and describes the biochemical and molecular properties of these enzymes, which are typically thermostable and optimally active at high temperatures.
Abstract: Enzymes synthesized by hyperthermophiles (bacteria and archaea with optimal growth temperatures of >80°C), also called hyperthermophilic enzymes, are typically thermostable (i.e., resistant to irreversible inactivation at high temperatures) and are optimally active at high temperatures. These enzymes share the same catalytic mechanisms with their mesophilic counterparts. When cloned and expressed in mesophilic hosts, hyperthermophilic enzymes usually retain their thermal properties, indicating that these properties are genetically encoded. Sequence alignments, amino acid content comparisons, crystal structure comparisons, and mutagenesis experiments indicate that hyperthermophilic enzymes are, indeed, very similar to their mesophilic homologues. No single mechanism is responsible for the remarkable stability of hyperthermophilic enzymes. Increased thermostability must be found, instead, in a small number of highly specific alterations that often do not obey any obvious traffic rules. After briefly discussing the diversity of hyperthermophilic organisms, this review concentrates on the remarkable thermostability of their enzymes. The biochemical and molecular properties of hyperthermophilic enzymes are described. Mechanisms responsible for protein inactivation are reviewed. The molecular mechanisms involved in protein thermostabilization are discussed, including ion pairs, hydrogen bonds, hydrophobic interactions, disulfide bridges, packing, decrease of the entropy of unfolding, and intersubunit interactions. Finally, current uses and potential applications of thermophilic and hyperthermophilic enzymes as research reagents and as catalysts for industrial processes are described.

1,937 citations

Journal ArticleDOI
01 May 2003
TL;DR: In this article, various pre-treatment options as well as enzymatic saccharification of lignocellulosic biomass to fermentable sugars are reviewed and the barriers, progress, and prospects of developing an environmentally benign bioprocess for large-scale conversion of hemicellulose to fuel ethanol, xylitol, 2,3-butanediol, and other value added fermentation products are highlighted.
Abstract: Various agricultural residues, such as corn fiber, corn stover, wheat straw, rice straw, and sugarcane bagasse, contain about 20–40% hemicellulose, the second most abundant polysaccharide in nature. The conversion of hemicellulose to fuels and chemicals is problematic. In this paper, various pretreatment options as well as enzymatic saccharification of lignocellulosic biomass to fermentable sugars is reviewed. Our research dealing with the pretreatment and enzymatic saccharification of corn fiber and development of novel and improved enzymes such as endo-xylanase, β-xylosidase, and α-l-arabinofuranosidase for hemicellulose bioconversion is described. The barriers, progress, and prospects of developing an environmentally benign bioprocess for large-scale conversion of hemicellulose to fuel ethanol, xylitol, 2,3-butanediol, and other value-added fermentation products are highlighted.

1,651 citations

Journal ArticleDOI
TL;DR: This review summarizes the current knowledge of the cell wall polysaccharide-degrading enzymes from aspergilli and the genes by which they are encoded and describes the enzymatic pathways followed by tailored modifications by using specific enzymes purified from these fungi.
Abstract: Degradation of plant cell wall polysaccharides is of major importance in the food and feed, beverage, textile, and paper and pulp industries, as well as in several other industrial production processes. Enzymatic degradation of these polymers has received attention for many years and is becoming a more and more attractive alternative to chemical and mechanical processes. Over the past 15 years, much progress has been made in elucidating the structural characteristics of these polysaccharides and in characterizing the enzymes involved in their degradation and the genes of biotechnologically relevant microorganisms encoding these enzymes. The members of the fungal genus Aspergillus are commonly used for the production of polysaccharide-degrading enzymes. This genus produces a wide spectrum of cell wall-degrading enzymes, allowing not only complete degradation of the polysaccharides but also tailored modifications by using specific enzymes purified from these fungi. This review summarizes our current knowledge of the cell wall polysaccharide-degrading enzymes from aspergilli and the genes by which they are encoded.

1,040 citations

Journal ArticleDOI
TL;DR: The need for such a process, the cellulases of clostridia, their presence in extracellular complexes or organelles (the cellulosomes), the binding of the cellulosome to cellulose and to the cell surface, cellulase genetics, regulation of their synthesis, cocultures, ethanol tolerance, and metabolic pathway engineering for maximizing ethanol yield are discussed.
Abstract: Biomass conversion to ethanol as a liquid fuel by the thermophilic and anaerobic clostridia offers a potential partial solution to the problem of the world's dependence on petroleum for energy. Coculture of a cellulolytic strain and a saccharolytic strain of Clostridium on agricultural resources, as well as on urban and industrial cellulosic wastes, is a promising approach to an alternate energy source from an economic viewpoint. This review discusses the need for such a process, the cellulases of clostridia, their presence in extracellular complexes or organelles (the cellulosomes), the binding of the cellulosomes to cellulose and to the cell surface, cellulase genetics, regulation of their synthesis, cocultures, ethanol tolerance, and metabolic pathway engineering for maximizing ethanol yield.

888 citations

Journal ArticleDOI
TL;DR: It is shown that values of DeltaGr(0) for many microbially mediated reactions are highly temperature dependent, and that adopting values determined at 25 degrees C for systems at elevated temperatures introduces significant and unnecessary errors.
Abstract: Thermophilic and hyperthermophilic Archaea and Bacteria have been isolated from marine hydrothermal systems, heated sediments, continental solfataras, hot springs, water heaters, and industrial waste They catalyze a tremendous array of widely varying metabolic processes As determined in the laboratory, electron donors in thermophilic and hyperthermophilic microbial redox reactions include H2, Fe(2+), H2S, S, S2O3(2-), S4O6(2-), sulfide minerals, CH4, various mono-, di-, and hydroxy-carboxylic acids, alcohols, amino acids, and complex organic substrates; electron acceptors include O2, Fe(3+), CO2, CO, NO3(-), NO2(-), NO, N2O, SO4(2-), SO3(2-), S2O3(2-), and S Although many assimilatory and dissimilatory metabolic reactions have been identified for these groups of microorganisms, little attention has been paid to the energetics of these reactions In this review, standard molal Gibbs free energies (DeltaGr(0)) as a function of temperature to 200 degrees C are tabulated for 370 organic and inorganic redox, disproportionation, dissociation, hydrolysis, and solubility reactions directly or indirectly involved in microbial metabolism To calculate values of DeltaGr(0) for these and countless other reactions, the apparent standard molal Gibbs free energies of formation (DeltaG(0)) at temperatures to 200 degrees C are given for 307 solids, liquids, gases, and aqueous solutes It is shown that values of DeltaGr(0) for many microbially mediated reactions are highly temperature dependent, and that adopting values determined at 25 degrees C for systems at elevated temperatures introduces significant and unnecessary errors The metabolic processes considered here involve compounds that belong to the following chemical systems: H-O, H-O-N, H-O-S, H-O-N-S, H-O-C(inorganic), H-O-C, H-O-N-C, H-O-S-C, H-O-N-S-C(amino acids), H-O-S-C-metals/minerals, and H-O-P For four metabolic reactions of particular interest in thermophily and hyperthermophily (knallgas reaction, anaerobic sulfur and nitrate reduction, and autotrophic methanogenesis), values of the overall Gibbs free energy (DeltaGr) as a function of temperature are calculated for a wide range of chemical compositions likely to be present in near-surface and deep hydrothermal and geothermal systems

678 citations