Stereoselective Bioreduction of Ethyl 3-Oxo-3-(2-Thienyl) Propanoate Using the Short-Chain Dehydrogenase/Reductase ChKRED12

( Zhi-qiang Ren ),( Yan Liu ),( Xiao-qiong Pei ),( Zhong-liu Wu ) 한국미생물 · 생명공학회 2019 Journal of microbiology and biotechnology Vol.29 No.11

Ethyl (S)-3-hydroxy-3-(2-thienyl) propanoate ((S)-HEES) acts as a key chiral intermediate for the blockbuster antidepressant drug duloxetine, which can be achieved via the stereoselective bioreduction of ethyl 3-oxo-3-(2-thienyl) propanoate (KEES) that contains a 3-oxoacyl structure. The sequences of the short-chain dehydrogenase/reductases from Chryseobacterium sp. CA49 were analyzed, and the putative 3-oxoacyl-acyl-carrier-protein reductase, ChKRED12, was able to stereoselectively catalyze the NADPH-dependent reduction to produce (S)-HEES. The reductase activity of ChKRED12 towards other substrates with 3- oxoacyl structure were confirmed with excellent stereoselectivity (>99% enantiomeric excess) in most cases. When coupled with a cofactor recycling system using glucose dehydrogenase, the ChKRED12 was able to catalyze the complete conversion of 100 g/l KEES within 12 h, yielding the enantiopure product with >99% ee, showing a remarkable potential to produce (S)-HEES.

Ciliogenesis is reciprocally regulated by PPARA and NR1H4/FXR through controlling autophagy in vitro and in vivo

Liu, Zhi-qiang,Lee, Joon No,Son, Myeongjoo,Lim, Jae-Young,Dutta, Raghbendra Kumar,Maharjan, Yunash,Kwak, SeongAe,Oh, Goo Taeg,Byun, Kyunghee,Choe, Seong-Kyu,Park, Raekil Landes Bioscience 2018 AUTOPHAGY Vol.14 No.6

<P>The primary cilia are evolutionarily conserved microtubule-based cellular organelles that perceive metabolic status and thus link the sensory system to cellular signaling pathways. Therefore, ciliogenesis is thought to be tightly linked to autophagy, which is also regulated by nutrient-sensing transcription factors, such as PPARA (peroxisome proliferator activated receptor alpha) and NR1H4/FXR (nuclear receptor subfamily 1, group H, member 4). However, the relationship between these factors and ciliogenesis has not been clearly demonstrated. Here, we present direct evidence for the involvement of macroautophagic/autophagic regulators in controlling ciliogenesis. We showed that activation of PPARA facilitated ciliogenesis independently of cellular nutritional states. Importantly, PPARA-induced ciliogenesis was mediated by controlling autophagy, since either pharmacological or genetic inactivation of autophagy significantly repressed ciliogenesis. Moreover, we showed that pharmacological activator of autophagy, rapamycin, recovered repressed ciliogenesis in ppara(-/-) cells. Conversely, activation of NR1H4 repressed cilia formation, while knockdown of NR1H4 enhanced ciliogenesis by inducing autophagy. The reciprocal activities of PPARA and NR1H4 in regulating ciliogenesis were highlighted in a condition where de-repressed ciliogenesis by NR1H4 knockdown was further enhanced by PPARA activation. The in vivo roles of PPARA and NR1H4 in regulating ciliogenesis were examined in greater detail in ppara(-/-) mice. In response to starvation, ciliogenesis was facilitated in wild-type mice via enhanced autophagy in kidney, while ppara(-/-) mice displayed impaired autophagy and kidney damage resembling ciliopathy. Furthermore, an NR1H4 agonist exacerbated kidney damage associated with starvation in ppara(-/-) mice. These findings indicate a previously unknown role for PPARA and NR1H4 in regulating the autophagy-ciliogenesis axis in vivo.</P>

Driving shaft fatigue optimization design of W type profile twin-screw pumps

Zhi-Jie Liu,Yu-Chong Zhao,Zhi-Qiang Gan,Dong-Lin Hui 대한기계학회 2018 JOURNAL OF MECHANICAL SCIENCE AND TECHNOLOGY Vol.32 No.11

Under changeable pumped medium and working environment, the twin-screw pump is prone to be broken by fatigue failures. A structure optimization design model and method of the driving shaft are presented based on response surface methodology and finite element analysis. In this model, the shaft diameter, chamfering degree and the shaft extension of the power end are selected as optimization variables, the limit values of the variables and maximal normal deformation of the spindle are considered as the constraint conditions, and the minimization of the equivalent alternating stress on the dangerous shaft section is taken as the optimization objective so as to improve the shaft fatigue reliability. The optimization results of a case show that the equivalent alternating stress on the dangerous spindle section reduces by 26.2 %, and the maximal normal deformation decreases by 25.2 % compared with the original design. In addition, the infinite life reliability and fatigue safety factors both meet the design requirements.

Statistical Optimization of Culture Media and Conditions for Production of Mannan by Saccharomyces cerevisiae

Hong-Zhi Liu,Qiang Wang,Yuan-Yuan Liu,Fang Fang 한국생물공학회 2009 Biotechnology and Bioprocess Engineering Vol.14 No.5

In view of the increase in Saccharomyces cerevisiae mannan content, the culture medium and condition for S.cerevisiae were optimized in this study. The influence of culture medium ingredients such as carbon and nitrogen sources, inorganic ion, and enzyme activator on mannan production were evaluated using factional design. The mathematical model was established by the quadratic rotary combination design through response surface analysis. The optimized concentrations of culture medium were determined as follows: 4.98 g/100 mL, sucrose; 4.39 g/100 mL, soybean peptone; 3.10 g/100 mL, yeast extract; and 2.21 g/100 mL, glycerol. The optimized culture medium increased mannan production from 82.7 ± 3.4 mg/100 mL to 162.53 ± 3.47 mg/100 mL. The influence of original pH, inoculum size, temperature, and media volume on mannan production was evaluated and confirmed by orthogonale experimental design, with the order of effect as follows: media volume > temperature > initial pH > inoculation size. The optimized culture condition was pH, 5; inoculum size, 5 ml; temperature, 32oC; and media volume, 40 mL. The maximum mannan production increased to 258.5 ± 9.1 mg/100 mL at the optimum culture condition.=It was evident that the mannan production was affected significantly by culture medium and condition optimization (é < 0.01). In view of the increase in Saccharomyces cerevisiae mannan content, the culture medium and condition for S.cerevisiae were optimized in this study. The influence of culture medium ingredients such as carbon and nitrogen sources, inorganic ion, and enzyme activator on mannan production were evaluated using factional design. The mathematical model was established by the quadratic rotary combination design through response surface analysis. The optimized concentrations of culture medium were determined as follows: 4.98 g/100 mL, sucrose; 4.39 g/100 mL, soybean peptone; 3.10 g/100 mL, yeast extract; and 2.21 g/100 mL, glycerol. The optimized culture medium increased mannan production from 82.7 ± 3.4 mg/100 mL to 162.53 ± 3.47 mg/100 mL. The influence of original pH, inoculum size, temperature, and media volume on mannan production was evaluated and confirmed by orthogonale experimental design, with the order of effect as follows: media volume > temperature > initial pH > inoculation size. The optimized culture condition was pH, 5; inoculum size, 5 ml; temperature, 32oC; and media volume, 40 mL. The maximum mannan production increased to 258.5 ± 9.1 mg/100 mL at the optimum culture condition.=It was evident that the mannan production was affected significantly by culture medium and condition optimization (é < 0.01).

Sex-Related Outcomes of Successful Drug-Coated Balloon Treatment in De Novo Coronary Artery Disease

Liu Kun,신은석,전은정,박영준,Scot Garg,김태현,손창배,최병주,Lin Hui,Song Lin Yuan,Wang Zhi,Jiang Hao,Shi Zhentao,Tang Qiang 연세대학교의과대학 2021 Yonsei medical journal Vol.62 No.11

Purpose: Although drug-coated balloon (DCB) treatment is known to be effective for de novo lesions, the influence of sex on angiographic and clinical outcomes remains unknown. This study aimed to investigate the angiographic and clinical impact of DCB treatment in patients with de novo coronary lesions according to sex. Materials and Methods: A total of 227 patients successfully treated with DCB were retrospectively enrolled and divided into two groups according to sex. The primary endpoint was late lumen loss (LLL) at 6-month angiography, and the secondary endpoint was target vessel failure (TVF), which included cardiac death, target vessel myocardial infarction, target lesion revascularization, and target vessel thrombosis. Results: The study enrolled 60 women (26.4%) and 167 men (73.6%). Compared to men, women had a smaller vessel size, larger DCB to reference vessel ratio, and more dissections after DCB treatment (55.0% vs. 37.1%, p=0.016). Women also had a significantly higher LLL compared to men (0.12±0.26 mm vs. 0.02±0.22 mm, p=0.012) at the 6-month follow-up angiography. During a median follow-up of 3.4 years (range 12.7–28.9 months), TVF was similar (women 6.7% vs. men 7.8%, p=0.944). In multivariable analysis, women were independently associated with a higher LLL. Conclusion: LLL was higher in women, but there was no difference in TVF between women and men. Based on multivariable analysis, the women sex was an independent predictor of higher LLL (Impact of Drug-coated Balloon Treatment in de Novo Coronary Lesion; NCT04619277).

Functions of Membrane-bound Alcohol Dehydrogenase and Aldehyde Dehydrogenase in the Bio-oxidation of Alcohols in Gluconobacter oxydans DSM 2003

Liu-Jing Wei,Ji-lai Zhou,Dan-ni Zhu,Bai-yi Cai,Jin-Ping Lin,Qiang Hua,Dong-Zhi Wei 한국생물공학회 2012 Biotechnology and Bioprocess Engineering Vol.17 No.6

In this study a new insight was provided to understand the functions of membrane-bound alcohol dehydrogenase (mADH) and aldehyde dehydrogenase (mALDH) in the bio-oxidation of primary alcohols, diols and poly alcohols using the resting cells of Gluconobacter oxydans DSM 2003 and its mutant strains as catalyst. The results demonstrated that though both mADH and mALDH participated in most of the oxidation of alcohols to their corresponding acid, the exact roles of these enzymes in each reaction might be different. For example,mADH played a key role in the oxidation of diols to its corresponding organic acid in G. oxydans, but it was dispensable when the primary alcohols were used as substrates. In contrast to mADH, mALDH appears to play a relatively minor role in organic acid-producing reactions because of the possible presence of other isoenzymes. Aldehydes were, however, found to be accumulated in the mALDH-deficient strain during the oxidation of alcohols.

Development and Evaluation of Loop-Mediated Isothermal Amplification Assay for Rapid Detection of Tylenchulus semipenetrans Using DNA Extracted from Soil

Zhi-Qiang Song,Ju-E Cheng,Fei-Xue Cheng,De-Yong Zhang,Yong Liu 한국식물병리학회 2017 Plant Pathology Journal Vol.33 No.2

Tylenchulus semipenetrans is an important and widespreadplant-parasitic nematode of citrus worldwideand can cause citrus slow decline disease leading tosignificant reduction in tree growth and yield. Rapidand accurate detection of T. semipenetrans in soil isimportant for the disease forecasting and management. In this study, a loop-mediated isothermal amplification(LAMP) assay was developed to detect T. semipenetrans using DNA extracted from soil. A setof five primers was designed from the internal transcribedspacer region (ITS1) of rDNA, and was highlyspecific to T. semipenetrans. The LAMP reaction wasperformed at 63°C for 60 min. The LAMP productwas visualized directly in one reaction tube by addingSYBR Green I. The detection limit of the LAMP assaywas 10–2 J2/0.5 g of soil, which was 10 times moresensitive than conventional PCR (10–1 J2/0.5 g of soil). Examination of 24 field soil samples revealed that theLAMP assay was applicable to a range of soils infestednaturally with T. semipenetrans, and the total assaytime was less than 2.5 h. These results indicated thatthe developed LAMP assay is a simple, rapid, sensitive,specific and accurate technique for detection of T. semipenetrans in field soil, and contributes to the effectivemanagement of citrus slow decline disease.

Differential expression of sex-related fat body proteins during the larval–pupal developmental stages of the silkworm (Bombyx mori)

Zhi-PingWu,Yan-Yan Liu,Guo-Qiang Chen,Ting-LiangWang,Jian-Zhong Tan 한국응용곤충학회 2014 Journal of Asia-Pacific Entomology Vol.17 No.1

The silkworm fat body is the site of many intermediary metabolic processes, and a source of sustenance forgrowth throughout the life cycle. Fat body proteins are responsible for storing nutrients, providing energy, andregulating hormones, and they have been identified using proteomic approaches. However, detailed differentialexpression of sex-related fat body proteins has not previously been evaluated. In the present study, we characterizedthe differential expression of sex-related fat body proteins, by using 2-dimensional gel electrophoresis(2-DE) followed bymass spectrometry identification and bioinformaticsmethods.We extracted the fat body proteinsfrom 5-day-old fifth instar larvae (L5), 10-day-old fifth instar larvae (corresponding to the end of spinning[LE]), and 0-day-old pupae (P0) of the multivoltine silkworm variety “Da Zao”. We confirmed the presence of 11important sex-specific expression proteins and 14 stage-specific expression proteins.We accurately identified 13of these specific expression proteins, including actin, calponin-like protein, 75 kDa subunit NADH, receptor foractivated protein kinase C from Bombyx mori (BmRACK), IMP (inosine monophosphate) cyclohydrolase, tropomyosin1, β-tubulin, hypothetical protein, antichymotrypsin precursor, and 30 K protein precursor.We showedthat BmRACK was differentially expressed betweenmale and female silkworms.Wediscuss the biological roles ofthe specific expression proteins during the larval–pupal developmental stages.

Development and Evaluation of Loop-Mediated Isothermal Amplification Assay for Rapid Detection of Tylenchulus semipenetrans Using DNA Extracted from Soil

Song, Zhi-Qiang,Cheng, Ju-E,Cheng, Fei-Xue,Zhang, De-Yong,Liu, Yong The Korean Society of Plant Pathology 2017 Plant Pathology Journal Vol.33 No.2

Tylenchulus semipenetrans is an important and widespread plant-parasitic nematode of citrus worldwide and can cause citrus slow decline disease leading to significant reduction in tree growth and yield. Rapid and accurate detection of T. semipenetrans in soil is important for the disease forecasting and management. In this study, a loop-mediated isothermal amplification (LAMP) assay was developed to detect T. semipenetrans using DNA extracted from soil. A set of five primers was designed from the internal transcribed spacer region (ITS1) of rDNA, and was highly specific to T. semipenetrans. The LAMP reaction was performed at $63^{\circ}C$ for 60 min. The LAMP product was visualized directly in one reaction tube by adding SYBR Green I. The detection limit of the LAMP assay was $10^{-2}J2/0.5g$ of soil, which was 10 times more sensitive than conventional PCR ($10^{-1}J2/0.5g$ of soil). Examination of 24 field soil samples revealed that the LAMP assay was applicable to a range of soils infested naturally with T. semipenetrans, and the total assay time was less than 2.5 h. These results indicated that the developed LAMP assay is a simple, rapid, sensitive, specific and accurate technique for detection of T. semipenetrans in field soil, and contributes to the effective management of citrus slow decline disease.

Down-regulation of Phosphoglucose Isomerase/Autocrine Motility Factor Enhances Gensenoside Rh2 Pharmacological Action on Leukemia KG1α Cells

You, Zhi-Mei,Zhao, Liang,Xia, Jing,Wei, Qiang,Liu, Yu-Min,Liu, Xiao-Yan,Chen, Di-Long,Li, Jing Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.3

Aims and Background: Ginsenoside Rh2, which exerts the potent anticancer action both in vitro and in vivo, is one of the most well characterized ginsenosides extracted from ginseng. Although its effects on cancer are significant, the underlying mechanisms remain unknown. In this study, we sought to elucidate possible links between ginsenoside Rh2 and phosphoglucose isomerase/autocrine motility factor (PGI/AMF). Methods: $KG1{\alpha}$, a leukemia cell line highly expressing PGI/AMF was assessed by western blot analysis and reverse transcription- PCR (RT-PCR) assay after transfection of a small interfering (si)-RNA to silence PGI/AMF. The effect of PGI/AMF on proliferation was measured by typan blue assay and antibody array. A cell counting kit (CCK)-8 and flow cytometry (FCM) were adopted to investigate the effects of Rh2 on PGI/AMF. The relationships between PGI/AMF and Rh2 associated with Akt, mTOR, Raptor, Rag were detected by western blot analysis. Results: KG1${\alpha}$ cells expressed PGI/AMF and its down-regulation significantly inhibited proliferation. The antibody array indicated that the probable mechanism was reduced expression of PARP, State1, SAPK/JNK and Erk1/2, while those of PRAS40 and p38 were up-regulated. Silencing of PGI/AMF enhanced the sensibility of $KG1{\alpha}$ to Rh2 by suppressing the expression of mTOR, Raptor and Akt. Conclusion: These results suggested that ginsenoside Rh2 suppressed the proliferation of $KG1{\alpha}$, the same as down-regulation of PGI/AMF. Down-regulation of PGI/AMF enhanced the pharmacological effects of ginsenoside Rh2 on KG1${\alpha}$ by reducing Akt/mTOR signaling.