Optically pure(R)-γ-and(R)-δ-lactones can be prepared by intramolecular cyclization of chiral hydroxy acids/esters reduced asymmetrically from γ-and δ-keto acids/esters using Saccharomyces cerevisiae(S.cerevisiae)...Optically pure(R)-γ-and(R)-δ-lactones can be prepared by intramolecular cyclization of chiral hydroxy acids/esters reduced asymmetrically from γ-and δ-keto acids/esters using Saccharomyces cerevisiae(S.cerevisiae) as a whole-cell biocatalyst.However,some of the enzymes catalyzing these reactions in S.cerevisiae are still unknown up to date.In this report,two carbonyl reductases,OdCRl and OdCR2,were successfully discovered,and cloned from S.cerevisiae using a genome-mining approach,and overexpressed in Escherichia coli(E.coli).Compared with OdCR1,OdCR2 can reduce 4-oxodecanoic acid and 5-oxodecanoic acid asymmetrically with higher stereoselectivity,generating(R)-γ-decalactone(99% ee) and(R)-δ-decalactone(98% ee) in 85% and 92%yields,respectively.This is the first report of native enzymes from S.cerevisiae for the enzymatic synthesis of chiral γ-and δ-lactones which is of wide uses in food and cosmetic industries.展开更多
Foxtail millet (Setaria italica L.) is a drought-tolerant millet crop of arid and semi-arid regions. Aldo-keto reductases (AKRs) are significant part of plant defence mechanism, having an ability to confer multiple st...Foxtail millet (Setaria italica L.) is a drought-tolerant millet crop of arid and semi-arid regions. Aldo-keto reductases (AKRs) are significant part of plant defence mechanism, having an ability to confer multiple stress tolerance. In this study, AKR1 gene expression was studied in roots and leaves of foxtail millet subjected to different regimes of PEG- and NaCl-stress for seven days. The quantitative Real-time PCR expression analysis in both root and leaves showed upregulation of AKR1 gene during PEG and salt stress. A close correlation exits between expression of AKR1 gene and the rate of lipid peroxidation along with the retardation of growth. Tissue-specific differences were found in the AKR1 gene expression to the stress intensities studied. The reduction in root and shoot growth under both stress conditions were dependent on stress severity. The level of lipid peroxidation as indicated by MDA formation was significantly increased in roots and leaves along with increased stress levels. Finally, these findings support the early responsive nature of AKR1 gene and seem to be associated at least in part with its ability to contribute in antioxidant defence related pathways which could provide a better protection against oxidative stress under stress conditions.展开更多
We screened 15 Agromyces strains from the Microbacteriaceae family and 16 Gordonia strains from the Gordoniaceae family to investigate their biocatalytic ability to reduce carbonyl compounds. Two Agromyces strains (A....We screened 15 Agromyces strains from the Microbacteriaceae family and 16 Gordonia strains from the Gordoniaceae family to investigate their biocatalytic ability to reduce carbonyl compounds. Two Agromyces strains (A. soli NBRC109063 and A. humatus NBRC109085) and two Gordonia strains (G. hydrophobica NBRC16057 and G. malaquae NBRC108250) grew well in 230 medium. The stereoselective reduction of various carbonyl compounds using these four strains was investigated. We discovered that these strains can reduce aliphatic and aromatic α-keto esters and an aromatic α-keto amide. On the basis of the conversion rate and stereoselectivity of the alcohols produced, G. hydrophobica NBRC16057 is a potential biocatalyst for the stereoselective reduction of α-keto esters and an aromatic α-keto amide to the corresponding chiral alcohols. Our results also suggest that the reduction of ethyl 2-methylacetoacetate by wet G. hydrophobica NBRC16057 cells in the presence of L-glutamate is useful for the production of chiral ethyl 3-hydroxy-2-methylbutanoate.展开更多
The\dDDe)Rhly --CU)Zjw(CO)3(jab--Cr,ilo,W)heterobinuclear m,t'1carbonyl c ompQunds were synthesized and characterized by ic and h Iljlh'Their catalytic activit ies for CO hydrogecation were determined.
为有效提升吸波材料的电磁吸收强度和有效吸收带宽,采用球磨法对球状Fe-Si-Cr进行扁平化处理,通过机械共混法将不同形貌Fe-Si-Cr和羰基铁粉(Carbonyl iron powder,CIP)复合,研究不同质量比对复合材料电磁参数和微波吸收性能的影响。实...为有效提升吸波材料的电磁吸收强度和有效吸收带宽,采用球磨法对球状Fe-Si-Cr进行扁平化处理,通过机械共混法将不同形貌Fe-Si-Cr和羰基铁粉(Carbonyl iron powder,CIP)复合,研究不同质量比对复合材料电磁参数和微波吸收性能的影响。实验结果表明,大长径比可以提高材料的介电常数和磁导率,并使反射率峰值向低频移动。片状Fe-Si-Cr和CIP的低频吸波性能优于球形粒子。实验制备的复合材料中,样品I的最大反射损耗为-45.92 dB,有效吸收带宽为1.5 GHz。样品L的最大反射损耗为-22.11 dB,有效吸收带宽大于6.2 GHz。吸波剂的形貌对材料的电磁吸收性能有显著影响。将不同形貌的CIP和Fe-Si-Cr按不同配比复合后,可以得到不同频率下性能优良的吸波体。利用阻抗匹配函数可以从理论上预测出反射损耗峰值对应的吸波剂厚度和频率。双层吸波材料相比于单层吸波材料,其有效吸收带宽更大,可以通过改变匹配层、吸收层的厚度来获得不同频率下吸波性能优良的吸波体,更易满足新型吸波材料“薄、轻、宽、强”的要求。展开更多
目的:探讨前列腺癌上调的长非编码RNA-1(prostate cancer up-regulated long non-coding RNA-1,PlncRNA-1)对低氧环境下胶质瘤相关的巨噬细胞浸润和极化的影响及其相关机制。方法:低氧诱导胶质瘤细胞U87,检测Control组和低氧Hypo组中Pln...目的:探讨前列腺癌上调的长非编码RNA-1(prostate cancer up-regulated long non-coding RNA-1,PlncRNA-1)对低氧环境下胶质瘤相关的巨噬细胞浸润和极化的影响及其相关机制。方法:低氧诱导胶质瘤细胞U87,检测Control组和低氧Hypo组中PlncRNA-1和羰基还原酶3(carbonyl reductase 3,CBR3)的表达。低氧的U87细胞分为4组,即U87细胞转染PlncRNA-1过表达质粒组(pcDNA-PlncRNA-1组)、空载体组(pcDNA-Empty组)、联合转染pcDNA-PlncRNA-1和CBR3的沉默寡核苷酸序列(si-CBR3)组(pcDNA-PlncRNA-1+si-CBR3组)及联合转染pcDNA-PlncRNA-1和si-CBR3的阴性对照组(pcDNA-PlncRNA-1+si-NC组),并分别与THP-1细胞在低氧环境下共培养。采用流式细胞术或者实时荧光定量PCR技术(real-timefluorescent quantitative PCR,RT-qPCR)法检测这4组中THP-1细胞的M1型标志物(CD163、IL-12、TNF-α)、M2型标志物(CD86、IL-10、趋化因子CCL-22)的变化;采用Transwell细胞迁移实验检测THP-1细胞迁移能力。结果:Hypo组PlncRNA-1和CBR3的表达水平均高于Control组(均P<0.05);pcDNA-PlncRNA-1组的CBR3表达水平明显高于pcDNA-Empty组(P<0.05)。对比pcDNA-Empty组,pcDNA-PlncRNA-1组胶质瘤细胞与THP-1细胞共培养促进THP-1细胞向M1表型转变,并且促进THP-1细胞迁移(均P<0.05)。而对比pcDNA-PlncRNA-1+si-NC组,pcDNA-PlncRNA-1+si-CBR3组抑制THP-1细胞向M1表型转变,并且抑制THP-1细胞迁移(均P<0.05)。结论:本研究证实低氧环境下PlncRNA-1通过上调CBR3促进胶质瘤相关巨噬细胞浸润和M1极化。展开更多
The bimetallic catalysts prepared from SiO_2-supported Ru-Co,Ru- Fe and Ru-Mo carbonyl clusters exhibited high yields and selectivities towards oxygenates such as C_1-C_5 from CO+H_2,in contrast to the catalysts prepa...The bimetallic catalysts prepared from SiO_2-supported Ru-Co,Ru- Fe and Ru-Mo carbonyl clusters exhibited high yields and selectivities towards oxygenates such as C_1-C_5 from CO+H_2,in contrast to the catalysts prepared from homometallic and bimetallic Ru,Ru-Ni,Ru-Rh,Ru-Mn,and Ru- Cr carbonyl clusters.The FTIR investigation revealed that the 1584 cm^(-1) species plays an important role in the formation of oxygenates in CO hydrogenation,which is possibly assigned to surface formyl species.展开更多
A novel nicotinamide adenine dinucleotide phosphate（NADPH）-dependent carbonyl reductase from Kluyverornyces marxianus（KmCR） was identified, which can convert various prochiral ketone esters and ketone substrates t...A novel nicotinamide adenine dinucleotide phosphate（NADPH）-dependent carbonyl reductase from Kluyverornyces marxianus（KmCR） was identified, which can convert various prochiral ketone esters and ketone substrates to their corresponding chiral alcohols. KmCR was over-expressed in E. coli BL21（DE3）, purified to homogeneity, and characterized. The purified enzyme exhibits the highest activity at 40℃ and pH=6.0. Based on the gel filtration and sodium dodecyl sulfate-polyacrylamide gel eiectrophoresis（SDS-PAGE） analysis, the monomeric protein was determined to have a molecular weight of approximate 39000. Vmax and Km of KmCR are 4.28 μmol.min^-1·mg^-1 and 0.41 mmol/L for ketone ester substrate ethyl 2-oxo-4-phenylbutyrate（OPBE）, 3.09μmol.min^-1·mg^-1 and 1.21 mmol/L for cofactor NADPH, respectively. Cofactor recycle was achieved by co-expression of KmCR and glucose dehydrogenase（GDH） in E. coli. Recombinant E. coli harboring KmCR and GDH showed moderate asymmetric reduction activity towards various α- and β-ketoesters, diaryl ketone substrates. In an aqueous/butyl acetate biphasic system, the whole-cell biocatalyst was used to prepare ethyl （R）-2-hydroxy-4- phenylbutanoate[（R）-HPBE] in an e.e. of 99.5% with a space-time yield of 433.6 g.L-1.d-1 and a yield of 80.3% at 270 g/L OPBE.展开更多
Prostaglandins （PGs） play a critical role in porcine reproduction, of which prostaglandin E2 （PGE2） and prostaglandin F2a （PGF2a） exert antiluteolytic and luteolysis actions, respectively. As a rate-limiting enz...Prostaglandins （PGs） play a critical role in porcine reproduction, of which prostaglandin E2 （PGE2） and prostaglandin F2a （PGF2a） exert antiluteolytic and luteolysis actions, respectively. As a rate-limiting enzyme, carbonyl reductase 1 （CBR1） catalyzes the conversion of PGE2 to PGF2a. A high ratio of PGE2：PGF2a is beneficial to the establishment and maintenance of porcine pregnancy. PG is essential for the establishment of pregnancy which resembles the proinflammatory response and nuclear factor KB （NF-KB） is involved in the process. Bioinformatic analysis has shown that NF-KB is a possible factor bound to two cis-regulatory elements in CBRI promoter. In this study, we cloned the 2997 bp （-2875/＋122） of the promoter, and constructed six 5＇-deleted dual-luciferase reporter recombinant vectors. In endometrial cells, the region of P2 （-16401＋7） exhibited the greatest transcriptional activity at driving luciferase expression, but not significantly different from that of P1 （-2089/＋7）. The activity of P1, P2, and P3 （-1019/＋7） was highly significantly higher than that of others （P〈0.01）, suggesting that two positive regulatory elements were likely present in the regions of -1640/-1019 and -1019/-647. The results also showed that the -1640/ -647 region was indispensable for the promoter. The results of chromatin immunoprecipitation （CHIP） demonstrated that the NF-KB subunit p65 binds to one site around -15451-1531. Using four reference genes, we found that the over-expression of p65 enhanced the expression of CBR1 （P〈0.05） in porcine endometrial epithelial cells, while knockdown of the p65 did not down-regulate the CBRI expression. These results indicated that NF-KB （p65） could bind to the special element of CBR1 gene promoter in porcine endometrial epithelial cells in vitro. The binding site of NF-KB was a positive regulator for the CBR1 gene promoter, but was not necessary for the basic expression.展开更多
基金financially sponsored by the National Key Research and Development Program of China (2016YFA0204300, 2019YFA09005000)the National Natural Science Foundation of China (21536004, 21776085, 21871085)+2 种基金the Natural Science Foundation of Shanghai (18ZR1409900)Key Project of the Shanghai Science and Technology Committee (18DZ1112703)the Fundamental Research Funds for the Central Universities (WF1714026)。
文摘Optically pure(R)-γ-and(R)-δ-lactones can be prepared by intramolecular cyclization of chiral hydroxy acids/esters reduced asymmetrically from γ-and δ-keto acids/esters using Saccharomyces cerevisiae(S.cerevisiae) as a whole-cell biocatalyst.However,some of the enzymes catalyzing these reactions in S.cerevisiae are still unknown up to date.In this report,two carbonyl reductases,OdCRl and OdCR2,were successfully discovered,and cloned from S.cerevisiae using a genome-mining approach,and overexpressed in Escherichia coli(E.coli).Compared with OdCR1,OdCR2 can reduce 4-oxodecanoic acid and 5-oxodecanoic acid asymmetrically with higher stereoselectivity,generating(R)-γ-decalactone(99% ee) and(R)-δ-decalactone(98% ee) in 85% and 92%yields,respectively.This is the first report of native enzymes from S.cerevisiae for the enzymatic synthesis of chiral γ-and δ-lactones which is of wide uses in food and cosmetic industries.
文摘Foxtail millet (Setaria italica L.) is a drought-tolerant millet crop of arid and semi-arid regions. Aldo-keto reductases (AKRs) are significant part of plant defence mechanism, having an ability to confer multiple stress tolerance. In this study, AKR1 gene expression was studied in roots and leaves of foxtail millet subjected to different regimes of PEG- and NaCl-stress for seven days. The quantitative Real-time PCR expression analysis in both root and leaves showed upregulation of AKR1 gene during PEG and salt stress. A close correlation exits between expression of AKR1 gene and the rate of lipid peroxidation along with the retardation of growth. Tissue-specific differences were found in the AKR1 gene expression to the stress intensities studied. The reduction in root and shoot growth under both stress conditions were dependent on stress severity. The level of lipid peroxidation as indicated by MDA formation was significantly increased in roots and leaves along with increased stress levels. Finally, these findings support the early responsive nature of AKR1 gene and seem to be associated at least in part with its ability to contribute in antioxidant defence related pathways which could provide a better protection against oxidative stress under stress conditions.
文摘We screened 15 Agromyces strains from the Microbacteriaceae family and 16 Gordonia strains from the Gordoniaceae family to investigate their biocatalytic ability to reduce carbonyl compounds. Two Agromyces strains (A. soli NBRC109063 and A. humatus NBRC109085) and two Gordonia strains (G. hydrophobica NBRC16057 and G. malaquae NBRC108250) grew well in 230 medium. The stereoselective reduction of various carbonyl compounds using these four strains was investigated. We discovered that these strains can reduce aliphatic and aromatic α-keto esters and an aromatic α-keto amide. On the basis of the conversion rate and stereoselectivity of the alcohols produced, G. hydrophobica NBRC16057 is a potential biocatalyst for the stereoselective reduction of α-keto esters and an aromatic α-keto amide to the corresponding chiral alcohols. Our results also suggest that the reduction of ethyl 2-methylacetoacetate by wet G. hydrophobica NBRC16057 cells in the presence of L-glutamate is useful for the production of chiral ethyl 3-hydroxy-2-methylbutanoate.
文摘The\dDDe)Rhly --CU)Zjw(CO)3(jab--Cr,ilo,W)heterobinuclear m,t'1carbonyl c ompQunds were synthesized and characterized by ic and h Iljlh'Their catalytic activit ies for CO hydrogecation were determined.
文摘目的:探讨前列腺癌上调的长非编码RNA-1(prostate cancer up-regulated long non-coding RNA-1,PlncRNA-1)对低氧环境下胶质瘤相关的巨噬细胞浸润和极化的影响及其相关机制。方法:低氧诱导胶质瘤细胞U87,检测Control组和低氧Hypo组中PlncRNA-1和羰基还原酶3(carbonyl reductase 3,CBR3)的表达。低氧的U87细胞分为4组,即U87细胞转染PlncRNA-1过表达质粒组(pcDNA-PlncRNA-1组)、空载体组(pcDNA-Empty组)、联合转染pcDNA-PlncRNA-1和CBR3的沉默寡核苷酸序列(si-CBR3)组(pcDNA-PlncRNA-1+si-CBR3组)及联合转染pcDNA-PlncRNA-1和si-CBR3的阴性对照组(pcDNA-PlncRNA-1+si-NC组),并分别与THP-1细胞在低氧环境下共培养。采用流式细胞术或者实时荧光定量PCR技术(real-timefluorescent quantitative PCR,RT-qPCR)法检测这4组中THP-1细胞的M1型标志物(CD163、IL-12、TNF-α)、M2型标志物(CD86、IL-10、趋化因子CCL-22)的变化;采用Transwell细胞迁移实验检测THP-1细胞迁移能力。结果:Hypo组PlncRNA-1和CBR3的表达水平均高于Control组(均P<0.05);pcDNA-PlncRNA-1组的CBR3表达水平明显高于pcDNA-Empty组(P<0.05)。对比pcDNA-Empty组,pcDNA-PlncRNA-1组胶质瘤细胞与THP-1细胞共培养促进THP-1细胞向M1表型转变,并且促进THP-1细胞迁移(均P<0.05)。而对比pcDNA-PlncRNA-1+si-NC组,pcDNA-PlncRNA-1+si-CBR3组抑制THP-1细胞向M1表型转变,并且抑制THP-1细胞迁移(均P<0.05)。结论:本研究证实低氧环境下PlncRNA-1通过上调CBR3促进胶质瘤相关巨噬细胞浸润和M1极化。
文摘The bimetallic catalysts prepared from SiO_2-supported Ru-Co,Ru- Fe and Ru-Mo carbonyl clusters exhibited high yields and selectivities towards oxygenates such as C_1-C_5 from CO+H_2,in contrast to the catalysts prepared from homometallic and bimetallic Ru,Ru-Ni,Ru-Rh,Ru-Mn,and Ru- Cr carbonyl clusters.The FTIR investigation revealed that the 1584 cm^(-1) species plays an important role in the formation of oxygenates in CO hydrogenation,which is possibly assigned to surface formyl species.
基金the National Basic Research and Development Program of China，the National Natural Science Foundation of China，the Project of New Century Excellent Talents in University of China，the Natural Science Foundation of Jiangsu Province,China，the Program of Introducing Talents of Discipline to Universities,China，the Priority Academic Program Development of Jiangsu Higher Education Institutions
文摘A novel nicotinamide adenine dinucleotide phosphate（NADPH）-dependent carbonyl reductase from Kluyverornyces marxianus（KmCR） was identified, which can convert various prochiral ketone esters and ketone substrates to their corresponding chiral alcohols. KmCR was over-expressed in E. coli BL21（DE3）, purified to homogeneity, and characterized. The purified enzyme exhibits the highest activity at 40℃ and pH=6.0. Based on the gel filtration and sodium dodecyl sulfate-polyacrylamide gel eiectrophoresis（SDS-PAGE） analysis, the monomeric protein was determined to have a molecular weight of approximate 39000. Vmax and Km of KmCR are 4.28 μmol.min^-1·mg^-1 and 0.41 mmol/L for ketone ester substrate ethyl 2-oxo-4-phenylbutyrate（OPBE）, 3.09μmol.min^-1·mg^-1 and 1.21 mmol/L for cofactor NADPH, respectively. Cofactor recycle was achieved by co-expression of KmCR and glucose dehydrogenase（GDH） in E. coli. Recombinant E. coli harboring KmCR and GDH showed moderate asymmetric reduction activity towards various α- and β-ketoesters, diaryl ketone substrates. In an aqueous/butyl acetate biphasic system, the whole-cell biocatalyst was used to prepare ethyl （R）-2-hydroxy-4- phenylbutanoate[（R）-HPBE] in an e.e. of 99.5% with a space-time yield of 433.6 g.L-1.d-1 and a yield of 80.3% at 270 g/L OPBE.
基金Project supported by the National Natural Science Foundation of China(No.31201771)the Earmarked Fund for China Agriculture Research System(No.CARS-36)
文摘Prostaglandins （PGs） play a critical role in porcine reproduction, of which prostaglandin E2 （PGE2） and prostaglandin F2a （PGF2a） exert antiluteolytic and luteolysis actions, respectively. As a rate-limiting enzyme, carbonyl reductase 1 （CBR1） catalyzes the conversion of PGE2 to PGF2a. A high ratio of PGE2：PGF2a is beneficial to the establishment and maintenance of porcine pregnancy. PG is essential for the establishment of pregnancy which resembles the proinflammatory response and nuclear factor KB （NF-KB） is involved in the process. Bioinformatic analysis has shown that NF-KB is a possible factor bound to two cis-regulatory elements in CBRI promoter. In this study, we cloned the 2997 bp （-2875/＋122） of the promoter, and constructed six 5＇-deleted dual-luciferase reporter recombinant vectors. In endometrial cells, the region of P2 （-16401＋7） exhibited the greatest transcriptional activity at driving luciferase expression, but not significantly different from that of P1 （-2089/＋7）. The activity of P1, P2, and P3 （-1019/＋7） was highly significantly higher than that of others （P〈0.01）, suggesting that two positive regulatory elements were likely present in the regions of -1640/-1019 and -1019/-647. The results also showed that the -1640/ -647 region was indispensable for the promoter. The results of chromatin immunoprecipitation （CHIP） demonstrated that the NF-KB subunit p65 binds to one site around -15451-1531. Using four reference genes, we found that the over-expression of p65 enhanced the expression of CBR1 （P〈0.05） in porcine endometrial epithelial cells, while knockdown of the p65 did not down-regulate the CBRI expression. These results indicated that NF-KB （p65） could bind to the special element of CBR1 gene promoter in porcine endometrial epithelial cells in vitro. The binding site of NF-KB was a positive regulator for the CBR1 gene promoter, but was not necessary for the basic expression.
