• The Microorganisms and Industry
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• v.18 no.3
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• pp.2-9
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• 1992

이 논문에서는 대사공학에의 응용에 필수적이며 또한 그 자체의 기술이 학문적으로 상당히 관심을 끄는 C. acetobutylicum에서의 primary metabolic gene cloning에 대하여 정리해 보고자 한다. 우선 C. acetobutylicum의 primary metabolism과 일반적인 대사 조절에 대하여 간략히 살펴보고 이에 관여한 효소들과 gene cloning에 대하여 기술하고자 한다.

• Journal of Microbiology and Biotechnology
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• v.26 no.4
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• pp.725-729
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• 2016

A novel genome-engineering tool in Clostridium acetobutylicum was developed based on single-crossover homologous recombination. A small-sized non-replicable plasmid, pHKO1, was designed for efficient integration into the C. acetobutylicum genome. The integrated pHKO1 plasmid backbone, which included an antibiotic resistance gene, can be excised in vivo by Flp recombinase, leaving a single flippase recognition target sequence in the middle of the targeted gene. Since the pSHL-FLP plasmid, the carrier of the Flp recombinase gene, employed the segregationally unstable pAMβ1 replicon, the plasmid was rapidly cured from the mutant C. acetobutylicum. Consequently, our method makes it easier to engineer C. acetobutylicum.

• Journal of Microbiology and Biotechnology
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• v.25 no.10
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• pp.1702-1708
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• 2015

We have developed a new shuttle plasmid, designated as pLK1-MCS that can replicate in both Clostridium acetobutylicum and Escherichia coli, by combining the pUB110 and pUC19 plasmids. Plasmid pLK1-MCS replicated more stably than previously reported plasmids containing either the pIM13 or the pAMβ1 replicon in the absence of antibiotic selective pressure. The transfer frequency of pLK1-MCS into C. acetobutylicum was similar to the transfer frequency of other shuttle plasmids. We complemented C. acetobutylicum ML1 (that does not produce solvents such as acetone, butanol, and ethanol owing to loss of the megaplasmid pSOL1 harboring the adhE1-ctfAB-adc operon) by introducing pLK1-MCS carrying the adhE1-ctfAB-adc operon into C. acetobutylicum ML1. The transformed cells were able to resume anaerobic solvent production, indicating that the new shuttle plasmid has the potential for practical use in microbial biotechnology.

• KSBB Journal
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• v.8 no.1
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• pp.1-9
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• 1993

The acetone-butanol fermentation by C. acetobutylicum has gained increasing attention for the following reasons. First, the finite supply of petrochemical resources, combined with increasing concern over global environmental effects and the unstable nature of the price of petroleum has renewed interest in the development of fermentation technology that allows utilzation of biomass wastes for the production of alcohol. Second, it serves as excellent model system for understading the regulation and molecular biology of tightly regulated complex primary metabolism, and for applications of metabolic engineering. In this review various aspects of acetone-butanol fermentation by C. acetobutylicm including strain and fermentation characteristics, enzyme regulation, and solvent formation mechanism, and product recovery and summarized.

• Korean Journal of Microbiology
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• v.28 no.3
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• pp.268-273
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• 1990

In order to elucidate the acid tolerance mechanism of Clostridium acetobutylicum against organic acid, the maintenance energy with added butyrate at different pH was determined. Maintenance coeffecient in acidogenic chemostat was higher at pH 6.5 than at pH 5.5 showing that this organism is an acidophile. The addition of butyrate at pH 5.5 and different dilution rate caused linear decrease of the cell concentration though $Y_{ATP}$ did not decrease with increasing undissociated organic acid. $Y_{ATP}$ decreased by increasing the concentration of undissociated organic acid at pH 5.0 by the addition of butyrate. From these results it is hypothesized that the ATP conaumption for pH stat of acidophile C. acetobutylicum is increased at the circumstance with over 30mM of undissociated organic acid.cid.

• Journal of Microbiology and Biotechnology
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• v.31 no.10
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• pp.1393-1400
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• 2021

Acetone-butanol-ethanol (ABE) fermentation by the anaerobic bacterium Clostridium acetobutylicum has been considered a promising process of industrial biofuel production. Phosphotransbutyrylase (phosphate butyryltransferase, PTB) plays a crucial role in butyrate metabolism by catalyzing the reversible conversion of butyryl-CoA into butyryl phosphate. Here, we report the crystal structure of PTB from the Clostridial host for ABE fermentation, C. acetobutylicum, (CaPTB) at a 2.9 Å resolution. The overall structure of the CaPTB monomer is quite similar to those of other acyltransferases, with some regional structural differences. The monomeric structure of CaPTB consists of two distinct domains, the N- and C-terminal domains. The active site cleft was formed at the interface between the two domains. Interestingly, the crystal structure of CaPTB contained eight molecules per asymmetric unit, forming an octamer, and the size-exclusion chromatography experiment also suggested that the enzyme exists as an octamer in solution. The structural analysis of CaPTB identifies the substrate binding mode of the enzyme and comparisons with other acyltransferase structures lead us to speculate that the enzyme undergoes a conformational change upon binding of its substrate.

• Microbiology and Biotechnology Letters
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• v.45 no.3
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• pp.236-242
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• 2017

The conversion of marine biomass to renewable energy has been considered an alternative to fossil fuels. Butanol, in particular, can be used directly as a fuel. In this experiment, the brown alga Undaria pinnatifida was selected as a biomass for biobutanol production. Hyper thermal (HT) acid hydrolysis was used as an acid hydrolysis method to produce monosaccharides. The optimal pretreatment conditions for U. pinnatifida were determined as slurry with 10% (w/v) U. pinnatifida content and 270 mM $H_2SO_4$, and heating at $160^{\circ}C$ for 7.5 min. Enzymatic saccharification was carried out with Celluclast 1.5 L, Viscozyme L, and Ultraflo Max. The optimal saccharification condition was 12 U/ml Viscozyme L. Fermentations were carried out for the production of acetone, butanol, and ethanol by Clostridium acetobutylicum KCTC 1724, Clostridium beijerinckii KCTC 1785, and Clostridium tyrobutyricum KCTC 5387. The fermentations were carried out using a pH-control. The optimal ABE fermentation condition determined using C. acetobutylicum KCTC 1724 adapted to 160 g/l mannitol. An ABE concentration of 9.05 g/l (0.99 g/l acetone, 5.62 g/l butanol, 2.44 g/l ethanol) was obtained by the consumption of 24.14 g/l monosaccharide with $Y_{ABE}$ of 0.37 in pH 5.0.

• Korean Journal of Microbiology
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• v.26 no.3
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• pp.278-282
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• 1988

Rice straw was used in the acetone-butanol fermentation by semultaneous saccharification and fermentation (SSF) using Clostridium acetobutylicum and cellulolytic enzyme. Over 230 mM of solvent was produced from alkali treated rice straw of from ball-milled microcrystalline cellulose whilst only acidic fermentation products were formed from ball-milled rice straw. From the results it is concluded that rice straw used in the study contained an inhibitor for the solventogenesis by the organism which is insoluble in water and some organic solvent and destroyed by alkaline treatment.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 1986.12a
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• pp.509.2-509
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• 1986

Owing to the growing interest in the production of fuels and chemicals from biomass the well-know butanol-acetone fermentation as carried out by Clostridium acetobutylicum has been intensely studied again in recent years. Several solvent-yielding fermentation processes were established which are operated by using batch cultures or continuous cultures. 1 could be shown that under conditions of phosphate limitation an asporogenous mutant of C. acetobutylicum establishes itself in a chemostat which produces the solvents continuously. Attempts have been made to change the butanol/acetone ratio in favor of butanol production. A corresponding shift of the product spectrum can be achieved by carbon monoxide addition to the head space of the fermentation (B.H. Kim et al., App. Envioron. Microbiol. 48, 764-770 1984) or by iron limitation. Progress has been made in understanding the mechanism underlying the shift from acid to solvent prodcction. Experimental results are in agreement with the view that intracellular accumulation of acetic and butyric acid results in a shortage of phosphate and coenzyme A. This shortage may serve then as signal for the synthesis of the enzymes involved in the formation of acetone and butanol.

• Journal of Microbiology and Biotechnology
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• v.24 no.9
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• pp.1178-1188
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• 2014

In this study, the yeast Pichia pastoris was genetically modified to assemble minicellulosomes on its cell surface by the heterologous expression of a truncated scaffoldin CipA from Clostridium acetobutylicum. Fluorescence microscopy and western blot analysis confirmed that CipA was targeted to the yeast cell surface and that NtEGD, the Nasutitermes takasagoensis endoglucanase that was fused with dockerin, interacted with CipA on the yeast cell surface, suggesting that the cohesin and dockerin domains and cellulose-binding module of C. acetobutylicum were functional in the yeasts. The enzymatic activities of the cellulases in the minicellulosomes that were displayed on the yeast cell surfaces increased dramatically following interaction with the cohesin-dockerin domains. Additionally, the hydrolysis efficiencies of NtEGD for carboxymethyl cellulose, microcrystal cellulose, and filter paper increased up to 1.4-fold, 2.0-fold, and 3.2-fold, respectively. To the best of our knowledge, this is the first report describing the expression of C. acetobutylicum minicellulosomes in yeast and the incorporation of animal cellulases into cellulosomes. This strategy of heterologous cellulase incorporation lends novel insight into the process of cellulosome assembly. Potentially, the surface display of cellulosomes, such as that reported in this study, may be utilized in the engineering of S. cerevisiae for ethanol production from cellulose and additional future applications.