Biosynthesis of Poly(2-hydroxybutyrate-co-lactate) in Metabolically Engineered Escherichia coli

채철기,이상엽,이혁,박시재,김유진,이세진,오영훈,양정은,주정찬,강경희,장영아,박아름,송봉근 한국생물공학회 2016 Biotechnology and Bioprocess Engineering Vol.21 No.1

We have previously reported in vivo biosynthesis of polyhydroxyalkanoates containing 2-hydroxyacid monomers such as lactate and 2-hydroxybutyrate in recombinant Escherichia coli strains by the expression of evolved Clostridium propionicum propionyl-CoA transferase (PctCp) and Pseudomonas sp. MBEL 6-19 polyhydroxyalkanoate (PHA) synthase 1 (PhaC1Ps6-19). Here, we report the biosynthesis of poly(2-hydroxybutyrate-co-lactate)[P(2HB-co- LA)] by direct fermentation of metabolically engineered E. coli strain. Among E. coli strains WL3110, XL1-Blue, and BL21(DE3), recombinant E. coli XL1-Blue strain expressing PhaC1437 and Pct540 produced P(76.4mol%2HB-co- 23.6mol%LA) to the highest content of 88 wt% when it was cultured in a chemically defined medium containing 20 g/L of glucose and 2 g/L of sodium 2-hydroxybutyrate. When recombinant E. coli XL1-Blue strain expressing PhaC1437 and Pct540 was cultured in a chemically defined medium containing 20 g/L of glucose and varying concentration of sodium 2-hydroxybutyrate, 2HB monomer fraction in P(2HB-co-LA) increased proportional to the concentration of sodium 2-hydroxybutyrate added to the culture medium. P(2HB-co-LA)] could also be produced from glucose as a sole carbon source without sodium 2-hydroxybutyrate into the culture medium. Recombinant E. coli XL1-Blue strain expressing the phaC1437, pct540, cimA3.7, and leuBCD genes together with the L. lactis Il1403 panE gene, successfully produced P(23.5mol%2HB-co- 76.5mol%LA)] to the polymer content of 19.4 wt% when it cultured in a chemically defined medium containing 20 g/L of glucose. The metabolic engineering strategy reported here should be useful for the production of novel copolymer P(2HB-co-LA)].

만성 췌장염 환자에서 췌관 스텐트 삽입 후 발생한 천공을 동반한 급성 담낭염 1예

김정욱,동석호,박철기,채정민,구자원,김태영,고원진,홍성훈 대한췌담도학회 2013 대한췌담도학회지 Vol.18 No.3

내시경 역행성 담췌관조영술과 관련된 급성 담낭염은 약 0.2-0.5% 정도로 보고되고 있으며 대부분이 담낭결석 이나, 조영제, 금속 스텐트 삽입과 관련이 있는 것으로 알 려져 있다. 하지만 ERCP 후 천공을 동반한 급성 담낭염은 아주 드물며 국내에서 진단적 ERCP 후 발생한 1예만 보고 하고 있다. 저자들은 만성 췌장염 환자에서 내시경적 치료 를 위해 췌관 스텐트 삽입 후 급성 담도염이 발생하였고 이에 대한 치료로 두 개의 플라스틱 담도 스텐트 삽입한 후 천공을 동반한 급성 담낭염을 경험하였다. 환자는 수술 없이 경피경간 담낭 배액술과 담도 스텐트 제거 후 증상 호전되어 퇴원할 수 있었다. 이에 저자들은 경험한 증례를 문헌 고찰과 함께 보고한다. Cholecystitis complicates approximately 0.2% to 0.5% of endoscopic retrograde cholangiopancreatographys (ERCPs). The risk appears to be correlated with the presence of stones in the gallbladder (GB), possibly filling of the GB with contrast, and placement of metal stents. However, acute acalculous cholecystitis with perforation after ERCP is extremely rare. A 56-year-old male with chronic pancreatitis was admitted for endoscopic treatment of chronic pancreatitis. Two days after pancreatic stent insertion, he presented with acute cholangitis without bile duct dilatation. He underwent two plastic biliary stents insertion. After two days, he had complained of acute right upper abdominal pain with muscle guarding. Abdominal computed tomographic scan showed focal discontinuity of GB wall and fluid collection of pericholecystic area and pelvic cavity. He had been successfully managed by percutaneous GB drainage and biliary stent removal without operation. Herein we report a case of acute acalculous cholecystitis with perforation that developed after ERCPs.

재조합 대장균에서 새로운 코엔자임 에이 트랜스퍼레이즈를 이용한 젖산을 모노머로 함유한 폴리하이드록시알칸산 생산 연구

김유진(You Jin Kim),채철기(Cheol Gi Chae),강경희(Kyoung Hee Kang),오영훈(Young Hoon Oh),주정찬(Jeong Chan Joo),송봉근(Bong Keun Song),이상엽(Sang Yup Lee),박시재(Si Jae Park) 한국생물공학회 2016 KSBB Journal Vol.31 No.1

Several CoA transferases from Clostridium beijerinckii, C. perfringens and Klebsiella pneumoniae were examined for biosynthesis of lactate-containing polyhydroxyalkanoates (PHAs) in recombinant Escherichia coli XL1-Blue strain. The CB3819 gene and the CB4543 gene from C. beijerinckii, the pct gene from C. perfringens and the pct gene from K. pneumoniae, which encodes putative CoA transferase gene, respectively, was co-expressed with the Pseudomonas sp. MBEL 6-19 phaC1437 gene encoding engineered Pseudomonas sp. MBEL 6-19 PHA synthase 1 (PhaC1Ps6-19) to examine its activity for the construction of key metabolic pathway to produce poly(3-hydroxybutyrate-co-lactate) [P(3HB-co-LA)]. The recombinant E. coli XL1-Blue expressing the phaC1437 gene and CB3819 gene synthesized poly(3-hydroxybutyrate) [P(3HB)] homopolymer to the P(3HB) content of 60.5 wt% when it was cultured in a chemically defined medium containing 20 g/L of glucose and 2 g/L of sodium 3-hydroxybutyrate. Expression of the phaC1437 gene and CB4543 gene in recombinant E. coli XL1-Blue also produced P(3HB) homopolymer to the P(3HB) content of 51.2 wt% in the same culture condition. Expression of the phaC1437 gene and the K. pneumoniae pct gene in recombinant E. coli XL1-Blue could not result in the production of PHAs in the same culture condition. However, the recombinant E. coli XL1-Blue expressing the phaC1437 gene and the C. perfringens gene could produce poly(3-hydroxybutyrate-co-lactate [P(86.4mol%3HB-co- 13.7 mol%LA) up to the PHA content of 10.6 wt% in the same culture condition. Newly examined CoA transfereases in this study may be useful for the construction of engineered E. coli strains to produce PHA containing novel monomer such lactate.

Screening of Microorganisms able to Degrade Low-rank Coal in Aerobic Conditions: Potential Coal Biosolubilization Mediators from Coal to Biochemicals

Yokimiko David,Mary Grace Baylon,Sudheer D. V. N. Pamidimarri,Kei-Anne Baritugo,채철기,김유진,김태완,김민식,나정걸,박시재 한국생물공학회 2017 Biotechnology and Bioprocess Engineering Vol.22 No.2

Coal is one of the major sources of energy, fuel, and other related chemicals. The processes to utilize coal for energy, fuel and other chemicals such as coal combustion, liquefaction, carbonization, and gasification pose a great threat to the environment by emitting toxic particles and CO2 to the atmosphere. Thus, biological beneficiation of coal can be a good strategy to utilize coal with environmental sustainability. Here, we report the screening of microorganisms able to degrade or depolymerize coal. These host strains are potential candidates for the development of biological treatment process of coal. A total of 45 microbial strains were isolated from sludge enriched with coal and were identified based on 16S rRNA sequencing. Four strains of three genera, Cupriavidus sp., Pseudomonas sp., and Alcaligenes sp., were further characterized for their abilities to degrade coal. The degree of coal degradation was analyzed by measuring the increase in absorbance at 450 nm by UV spectroscopy. These microorganisms were also able to increase the pH of the culture media as a response to the acidic nature of coal. Laccase-like activity was also found in these strains when tested for RBBR dye degradation. Since biological degradation of coal through the use of microorganisms is a good alternative to chemical combustion of coal, microbial strains isolated in this study can be potential biological catalysts for coal conversion into valuable chemicals.

Bio-solubilization of the untreated low rank coal by alkali-producing bacteria isolated from soil

메리 그레이스 베일런,요키미코 데이비드,Sudheer D. V. N. Pamidimarri,Kei-Anne Baritugo,채철기,김유진,김태원,김민식,나정걸,박시재 한국화학공학회 2017 Korean Journal of Chemical Engineering Vol.34 No.1

Coal is a hydrocarbon-rich fossil fuel considered as a possible replacement for petroleum as a feedstock for the production of fuel and valuable chemicals. In this study, bacteria capable of solubilizing untreated low rank coal were isolated from soil. A total of 19 microorganisms were isolated from soil enriched in MR medium with coal and were identified based on 16S rRNA sequencing. The identified soil isolates belonging to the genera Citricoccus, Comamonas, Cupriavidus, Sphingomonas, and Sphingopyxis were screened based on their growth in the chemically defined MR medium containing different concentrations of coal. Among the identified microbial strains, Cupriavidus necator S2A2, Sphingopyxis ginsengisoli S2B14 and Sphingomonas sp. S2B18 were further characterized for their ability to degrade low-rank coal. Cupriavidus necator S2A2, Sphingopyxis ginsengisoli S2B14 and Sphingomonas sp. S2B18 were found to solubilize untreated low-rank coal as indicated by the release of solubilized coal products detected at OD450 when they were grown in LB medium containing 1% coal. Sphingomonas sp. S2B18 showed the highest coal solubilization activity, based on the high absorbance of its culture supernatant (0.190). Although laccase-like activity was not detected in these strains when tested for RBBR dye degradation, increase in the pH of the culture medium up to 8.25- 8.34 was observed. This may be attributed to the excretion of alkaline substances in the culture medium. Since biosolubilization of coal by microorganisms is a good alternative for the chemical conversion of coal, microorganisms screened in this study can be potentially used as biological catalysts for the conversion of coal into valuable chemicals.