β-Poly(L-malic acid)(PMLA)is a water-soluble biopolymer used in food,medicine and other industries.To date,the biosynthesis pathway of PMLA has not been fully elucidated.In this study,we sequenced the transcriptom e ...β-Poly(L-malic acid)(PMLA)is a water-soluble biopolymer used in food,medicine and other industries.To date,the biosynthesis pathway of PMLA has not been fully elucidated.In this study,we sequenced the transcriptom e of strain Aureobasidium melanogenum under 20 g/L CaCO_(3) addition.The resulting sequencing reads were assembled and annotated for the differentially expressed genes(DEGs)analysis and novel transcripts identification.The result indicated that with the CaCO_(3) addition,the tricarboxylic cycle(TCA)cycle and glyoxylate pathway were up-regulated,and it also found that a non-ribosomal peptide synthetase(NRPS)like protein was highly expressed.The DEGs analysis showed a high expression level of malate dehydrogenase(MDHC)and phosphoenolpyruvate carboxykinase(PCKA)in the CaCO_(3) group,which indicated a cytosolic malate activity.We speculated that the malate should be transported to or synthesized in the cytoplasm,which was then polymerized to PMLA by the NRPS-like protein,accompanied by the up-regulated TCA cycle providing ATP for the polymerization.Depending on the analysis,we assumed that an NRPS-like protein,the TCA cycle,and the cytosolic malate together are contributing to the PMLA biosynthesis.展开更多
The marine yeast strain N13d, producing an extracellular amylase, was isolated from the deep sea sediments of the Pa-cific Ocean. This strain was identified to be Aureobasidium pullulans by 18S rRNA gene sequence anal...The marine yeast strain N13d, producing an extracellular amylase, was isolated from the deep sea sediments of the Pa-cific Ocean. This strain was identified to be Aureobasidium pullulans by 18S rRNA gene sequence analysis and routine yeast identi-fication methods. The optimal sea water medium for amylase production by this yeast strain was 1.0% peptone and 1.0% soluble starch with pH 4.0. The optimal conditions for amylase production by this yeast strain were with temperature 28 ℃, aeration rate 6 Lmin-1 and agitation speed 250 rmin-1. Under these conditions, 58.5 units of amylase activity per mg protein were produced within 56 h of fermentation.展开更多
The microbial production of either ester/lactones or enantio-enriched alcohols through Baeyer-Villiger oxidation or stereoselective reduction of ketones,respectively,is possible by using whole cells of A.subglaciale F...The microbial production of either ester/lactones or enantio-enriched alcohols through Baeyer-Villiger oxidation or stereoselective reduction of ketones,respectively,is possible by using whole cells of A.subglaciale F134 as a bifunctional biocatalyst.The chemoselective pattern of acetophenone biotransformation catalyzed by these cells can be regulated through reaction temperature,directing the reaction either towards oxidation or reduction products.The Baeyer–Villiger oxidation activity of A.subglaciale F134 whole cells is particularly dependent on reaction temperature.Acetophenone was transformed efficiently to phenol via the primary Baeyer–Villiger product phenyl acetate at 20℃ after 48 h with 100% conversion.In contrast,at 35℃,enantio-enriched(S)-1-phenylethanol was obtained as the sole product with 64% conversion and 89% ee.In addition,A.subglaciale F134 cells also catalyze the selective reduction of various structurally different aldehydes and ketones to alcohols with 40% to 100% yield,indicating broad substrate spectrum and good enantioselectivity in relevant cases.Our study provides a bifunctional biocatalyst systemthat can be used in Baeyer–Villiger oxidation aswell as in asymmetric carbonyl reduction,setting the stage for future work concerning the identification and isolation of the respective enzymes.展开更多
In the current study, in order to change the permeability of cell membrane and solve the problem of linked group of fungi mycelium, the method of adjusting osmotic pressure of medium and adding tween-80 was establishe...In the current study, in order to change the permeability of cell membrane and solve the problem of linked group of fungi mycelium, the method of adjusting osmotic pressure of medium and adding tween-80 was established. The utilized strain with relatively high exopolysaccharide (EPS) yield and low pigment level was obtained after the rejuvenation and sifting of long-preserved Aureobasidium pullulans strain. The optimal proportion of substrate was determined by means of orthogonal test. The transformation ratio of EPS was increased by 10% - 20% and the pigment content was greatly reduced. The fermenting liquor is between creamy white and pale yellow, and the white primary product can be gained without decolourization step. Furthermore, to magnify to 5 L bioreactor can get the similar result.展开更多
We have reported that A. pullulans 98 produces a high yield of cellulase. In this study, a carboxymethyl cellulase(CMCase) in the supernatant of the culture of A. pullulans 98 was purified to homogeneity, and the maxi...We have reported that A. pullulans 98 produces a high yield of cellulase. In this study, a carboxymethyl cellulase(CMCase) in the supernatant of the culture of A. pullulans 98 was purified to homogeneity, and the maximum production of CMCase was 4.51 U(mg protein)-1. The SDS-PAGE analysis showed that the molecular mass of the purified CMCase was 67.0 k Da. The optimal temperature of the purified enzyme with considerable thermosensitivity was 40℃, much lower than that of the CMCases from other fungi. The optimal p H of the enzyme was 5.6, and the activity profile was stable in a range of acidity(p H 5.0–6.0). The enzyme was activated by Na+, Mg2+, Ca2+, K+, Fe2+ and Cu2+, however, it was inhibited by Fe3+, Ba2+, Zn2+, Mn2+ and Ag+. Km and Vmax values of the purified enzyme were 4.7 mg m L-1 and 0.57 μmol L-1 min-1(mg protein)-1, respectively. Only oligosaccharides with different sizes were released from carboxymethylcellulose(CMC) after hydrolysis with the purified CMCase. The putative gene encoding CMCase was cloned from A. pullulans 98, which contained an open reading frame of 954 bp(EU978473). The protein deduced contained the conserved domain of cellulase superfamily(glucosyl hydrolase family 5). The N-terminal amino acid sequence of the purified CMCase was M-A-P-H-A-E-P-Q-S-Q-T-T-E-Q-T-S-S-G-Q-F, which was consistent with that deduced from the cloned gene. This suggested that the purified CMCase was indeed encoded by the cloned CMCase gene in this yeast.展开更多
Aureobasidium pullulans, a biocontrol agent for the annual weed Galium aparine L. was evaluated in vitro for its compatibility with commercial formulation of five herbicides at 1X(recommended field rate), 0.5X, 0.2X, ...Aureobasidium pullulans, a biocontrol agent for the annual weed Galium aparine L. was evaluated in vitro for its compatibility with commercial formulation of five herbicides at 1X(recommended field rate), 0.5X, 0.2X, 0.1X, 0.067 X, and 0.05 X concentrations. Germination of A. pullulans with paraquat, 2, 4-D, quizalofop-p, and clethodim treatment appeared reduced compared with germination of A. pullulans with fluroxypyr treatment at all concentrations. Stunted and shorter germ tubes in comparison with the control were observed with 2, 4-D, quizalofop-p, and clethodim at 0.2X. All concentration of paraquat, 2, 4-D, quizalofop-p, and clethodim except0.05 X, significantly decreased radial growth of A. pullulans compared with its growth on the untreated PDA medium. Field trials to further develop A. pullulans as biocontrol agent for control G. aparine L. was conducted to test the effectiveness of this fungus in wheat plots for 2 years at the same location in Xining. Treatments included spore suspensions of A. pullulans alone, a mixture of both fungus and fluroxypyr in wheat. Biocontrol agent effectiveness was estimated at approximately 7and 14 days after treatment, as disease incidence, percent weed control, and weed biomass reduction. Significant reduction in weed biomass occurred in combination treatments, and potential exists to tank mix A. pullulans with fluroxypyr. Leaf surface moisture and air temperatures following application may account for inconsistencies in field results between years. This fungal organisms show potential as bioherbicides for weeds in G. aparine L.展开更多
基金the financial support of the Tianjin Municipal Science and Technology Commission(17PTGCCX00190,17PTSYJC00080,17YFCZZC00310,and 16YFXTSF00460)the Tianjin Engineering Research Center of Microbial Metabolism and Fermentation Process Control(ZXKF20180301).
文摘β-Poly(L-malic acid)(PMLA)is a water-soluble biopolymer used in food,medicine and other industries.To date,the biosynthesis pathway of PMLA has not been fully elucidated.In this study,we sequenced the transcriptom e of strain Aureobasidium melanogenum under 20 g/L CaCO_(3) addition.The resulting sequencing reads were assembled and annotated for the differentially expressed genes(DEGs)analysis and novel transcripts identification.The result indicated that with the CaCO_(3) addition,the tricarboxylic cycle(TCA)cycle and glyoxylate pathway were up-regulated,and it also found that a non-ribosomal peptide synthetase(NRPS)like protein was highly expressed.The DEGs analysis showed a high expression level of malate dehydrogenase(MDHC)and phosphoenolpyruvate carboxykinase(PCKA)in the CaCO_(3) group,which indicated a cytosolic malate activity.We speculated that the malate should be transported to or synthesized in the cytoplasm,which was then polymerized to PMLA by the NRPS-like protein,accompanied by the up-regulated TCA cycle providing ATP for the polymerization.Depending on the analysis,we assumed that an NRPS-like protein,the TCA cycle,and the cytosolic malate together are contributing to the PMLA biosynthesis.
文摘The marine yeast strain N13d, producing an extracellular amylase, was isolated from the deep sea sediments of the Pa-cific Ocean. This strain was identified to be Aureobasidium pullulans by 18S rRNA gene sequence analysis and routine yeast identi-fication methods. The optimal sea water medium for amylase production by this yeast strain was 1.0% peptone and 1.0% soluble starch with pH 4.0. The optimal conditions for amylase production by this yeast strain were with temperature 28 ℃, aeration rate 6 Lmin-1 and agitation speed 250 rmin-1. Under these conditions, 58.5 units of amylase activity per mg protein were produced within 56 h of fermentation.
基金financially supported by the National Natural Science Foundation of China(No.21646014 and 21776134)the program of Jiangsu Synergetic Innovation Center for Advanced Bio-Manufacture(XTE1851).
文摘The microbial production of either ester/lactones or enantio-enriched alcohols through Baeyer-Villiger oxidation or stereoselective reduction of ketones,respectively,is possible by using whole cells of A.subglaciale F134 as a bifunctional biocatalyst.The chemoselective pattern of acetophenone biotransformation catalyzed by these cells can be regulated through reaction temperature,directing the reaction either towards oxidation or reduction products.The Baeyer–Villiger oxidation activity of A.subglaciale F134 whole cells is particularly dependent on reaction temperature.Acetophenone was transformed efficiently to phenol via the primary Baeyer–Villiger product phenyl acetate at 20℃ after 48 h with 100% conversion.In contrast,at 35℃,enantio-enriched(S)-1-phenylethanol was obtained as the sole product with 64% conversion and 89% ee.In addition,A.subglaciale F134 cells also catalyze the selective reduction of various structurally different aldehydes and ketones to alcohols with 40% to 100% yield,indicating broad substrate spectrum and good enantioselectivity in relevant cases.Our study provides a bifunctional biocatalyst systemthat can be used in Baeyer–Villiger oxidation aswell as in asymmetric carbonyl reduction,setting the stage for future work concerning the identification and isolation of the respective enzymes.
文摘In the current study, in order to change the permeability of cell membrane and solve the problem of linked group of fungi mycelium, the method of adjusting osmotic pressure of medium and adding tween-80 was established. The utilized strain with relatively high exopolysaccharide (EPS) yield and low pigment level was obtained after the rejuvenation and sifting of long-preserved Aureobasidium pullulans strain. The optimal proportion of substrate was determined by means of orthogonal test. The transformation ratio of EPS was increased by 10% - 20% and the pigment content was greatly reduced. The fermenting liquor is between creamy white and pale yellow, and the white primary product can be gained without decolourization step. Furthermore, to magnify to 5 L bioreactor can get the similar result.
基金Qingdao Municipal Science and Technology Commission,Qingdao,China for providing financial support to this work(06-2-2-22-jch)
文摘We have reported that A. pullulans 98 produces a high yield of cellulase. In this study, a carboxymethyl cellulase(CMCase) in the supernatant of the culture of A. pullulans 98 was purified to homogeneity, and the maximum production of CMCase was 4.51 U(mg protein)-1. The SDS-PAGE analysis showed that the molecular mass of the purified CMCase was 67.0 k Da. The optimal temperature of the purified enzyme with considerable thermosensitivity was 40℃, much lower than that of the CMCases from other fungi. The optimal p H of the enzyme was 5.6, and the activity profile was stable in a range of acidity(p H 5.0–6.0). The enzyme was activated by Na+, Mg2+, Ca2+, K+, Fe2+ and Cu2+, however, it was inhibited by Fe3+, Ba2+, Zn2+, Mn2+ and Ag+. Km and Vmax values of the purified enzyme were 4.7 mg m L-1 and 0.57 μmol L-1 min-1(mg protein)-1, respectively. Only oligosaccharides with different sizes were released from carboxymethylcellulose(CMC) after hydrolysis with the purified CMCase. The putative gene encoding CMCase was cloned from A. pullulans 98, which contained an open reading frame of 954 bp(EU978473). The protein deduced contained the conserved domain of cellulase superfamily(glucosyl hydrolase family 5). The N-terminal amino acid sequence of the purified CMCase was M-A-P-H-A-E-P-Q-S-Q-T-T-E-Q-T-S-S-G-Q-F, which was consistent with that deduced from the cloned gene. This suggested that the purified CMCase was indeed encoded by the cloned CMCase gene in this yeast.
基金Supported by National Natural Science Foundation of China(No.31160371,30860165)the National Key Technology R&D program of China(No.2012BAD19B02)the National High Technology Research and Development Program(863Program)of China(No.2011AA10A206)
文摘Aureobasidium pullulans, a biocontrol agent for the annual weed Galium aparine L. was evaluated in vitro for its compatibility with commercial formulation of five herbicides at 1X(recommended field rate), 0.5X, 0.2X, 0.1X, 0.067 X, and 0.05 X concentrations. Germination of A. pullulans with paraquat, 2, 4-D, quizalofop-p, and clethodim treatment appeared reduced compared with germination of A. pullulans with fluroxypyr treatment at all concentrations. Stunted and shorter germ tubes in comparison with the control were observed with 2, 4-D, quizalofop-p, and clethodim at 0.2X. All concentration of paraquat, 2, 4-D, quizalofop-p, and clethodim except0.05 X, significantly decreased radial growth of A. pullulans compared with its growth on the untreated PDA medium. Field trials to further develop A. pullulans as biocontrol agent for control G. aparine L. was conducted to test the effectiveness of this fungus in wheat plots for 2 years at the same location in Xining. Treatments included spore suspensions of A. pullulans alone, a mixture of both fungus and fluroxypyr in wheat. Biocontrol agent effectiveness was estimated at approximately 7and 14 days after treatment, as disease incidence, percent weed control, and weed biomass reduction. Significant reduction in weed biomass occurred in combination treatments, and potential exists to tank mix A. pullulans with fluroxypyr. Leaf surface moisture and air temperatures following application may account for inconsistencies in field results between years. This fungal organisms show potential as bioherbicides for weeds in G. aparine L.
