木糖酸氧化途径在枯草芽孢杆菌的构建及转化乙醇酸的研究
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摘要
通过过表达外源基因,旨在获得一株生产乙醇酸的食品安全菌。通过构建质粒在枯草芽孢杆菌A164S中过表达来自大肠杆菌的木糖酸降解途径,并用高效液相色谱对产物进行检测。诱导质粒表达后,重组枯草芽孢杆菌菌株成功地将木糖酸转化为乙醇酸,摩尔转化率达到42.54%。同时,枯草芽孢杆菌自身因存在非特异的醛缩酶及乙二醛脱氢酶,能够在只表达木糖酸脱水酶YjhG的情况下获得生物转化木糖酸的能力。成功构建了一株可利用木糖酸生产乙醇酸的枯草芽孢杆菌,并发现了可能的YjhH同工酶的存在。
It is aimed that a strain of a food-safety bacterium producing glycolic acid can be obtained by over-expressing foreign genes.An over-expression system comprising of xylonate degradation pathway from Escherichia coli was constructed and introduced into Bacillus subtilis(A164 S),glycolic acid was detected as the main product in the broth of recombinant B. subtilis,using high performance liquid chromatography(HPLC),and the conversion rate of xylonite to glycolic acid reached 42.54%. Meanwhile,Bacillus subtilis,because of its own nonspecific aldolase and glyoxal dehydrogenase,obtained the ability to biologically transform xylic acid under the condition that only xylose dehydratase YjhG was expressed. In conclusion,a strain of B. subtilis producing glycolic acid from xylic acid is successfully constructed and the possible existence of YjhH isozyme is uncovered.
It is aimed that a strain of a food-safety bacterium producing glycolic acid can be obtained by over-expressing foreign genes.An over-expression system comprising of xylonate degradation pathway from Escherichia coli was constructed and introduced into Bacillus subtilis(A164 S),glycolic acid was detected as the main product in the broth of recombinant B. subtilis,using high performance liquid chromatography(HPLC),and the conversion rate of xylonite to glycolic acid reached 42.54%. Meanwhile,Bacillus subtilis,because of its own nonspecific aldolase and glyoxal dehydrogenase,obtained the ability to biologically transform xylic acid under the condition that only xylose dehydratase YjhG was expressed. In conclusion,a strain of B. subtilis producing glycolic acid from xylic acid is successfully constructed and the possible existence of YjhH isozyme is uncovered.
引文
[1]Kabir SMM, Koh J. Effect of chelating agent in disperse dye dyeing on polyester fabric[J]. Fibers and Polymers, 2017, 18(12):2315-2321.
[2] Babilas P, Knie U, Abels C. Cosmetic and dermatologic use of alpha hydroxy acids[J]. J Dtsch Dermatol Ges, 2012, 10(7):488-491.
[3]Rendon MI, Berson DS, Cohen JL, et al. Evidence and considerations in the application of chemical peels in skin disorders and aesthetic resurfacing[J]. J Clin Aesthet Dermatol, 2010, 3(7):32.
[4]Hua X, Cao R, Zhou X, et al. Integrated process for scalable bioproduction of glycolic acid from cell catalysis of ethylene glycol[J]. Bioresource Technology, 2018, 268:402-407.
[5]Cabulong RB, Lee WK, Ba?ares AB, et al. Engineering Escherichia coli for glycolic acid production from D-xylose through the Dahms pathway and glyoxylate bypass[J]. Applied Microbiology and Biotechnology, 2018, 102(5):2179-2189.
[6]Alkim C, Trichez D, et al. The synthetic xylulose-1 phosphate pathway increases production of glycolic acid from xylose-rich sugar mixtures[J]. Biotechnology for Biofuels, 2016, 9(1):201.
[7]李宇展,刘伟,顾登平.草酸电还原研制乙醇酸[J].河北师范大学学报, 2003(4):385-387.
[8]Dahms AS. 3-Deoxy-D-pentulosonic acid aldolase and its role in a new pathway of D-xylose degradation[J]. Biochemical and biophysical research communications, 1974, 60(4):1433-1439.
[9]van Dijl JM, Hecker M. Bacillus subtilis:from soil bacterium to super-secreting cell factory[J]. Microb Cell Fact, 2013, 12:3.
[10]T?nnler S, Zamboni N, Kiraly C, et al. Screening of Bacillus subtilis transposon mutants with altered riboflavin production[J].Metabolic engineering, 2008, 10(5):216-226.
[11]李思杰,纪明华,赵利超,史吉平,孙俊松. N-乙酰神经氨酸合成途径在枯草芽孢杆菌的构建[J].食品工业科技, 2017,38(16):131-135.
[12]Liu H, Ramos KRM, Valdehuesa KNG, et al. Biosynthesis of ethylene glycol in Escherichia coli[J]. Applied Microbiology and Biotechnology, 2013, 97(8):3409-3417.
[13]Tai YS, Xiong M, Jambunathan P, et al. Engineering nonphosphorylative metabolism to generate lignocellulose-derived products[J]. Nature Chemical Biology, 2016, 12(4):247.
[14]Kolodkin-Gal I, Romero D, Cao S, et al. D-amino acids trigger biofilm disassembly[J]. Science, 2010, 328(5978):627-629.
[15]Cheng YL, Kalman LV, Kaiser D. The dsg gene of Myxococcus xanthus encodes a protein similar to translation initiation factor IF3[J]. Journal of Bacteriology, 1994, 176(5):1427-1433.
[16]Weimberg R. Pentose oxidation by Pseudomonas fragi[J]. J Biol Chem, 1961, 236(3):629-635.
[17]Meijnen JP, de Winde J H, et al. Establishment of oxidative D-xylose metabolism in Pseudomonas putida S12[J]. Applied and Environmental Microbiology, 2009, 75(9):2784-2791.
[2] Babilas P, Knie U, Abels C. Cosmetic and dermatologic use of alpha hydroxy acids[J]. J Dtsch Dermatol Ges, 2012, 10(7):488-491.
[3]Rendon MI, Berson DS, Cohen JL, et al. Evidence and considerations in the application of chemical peels in skin disorders and aesthetic resurfacing[J]. J Clin Aesthet Dermatol, 2010, 3(7):32.
[4]Hua X, Cao R, Zhou X, et al. Integrated process for scalable bioproduction of glycolic acid from cell catalysis of ethylene glycol[J]. Bioresource Technology, 2018, 268:402-407.
[5]Cabulong RB, Lee WK, Ba?ares AB, et al. Engineering Escherichia coli for glycolic acid production from D-xylose through the Dahms pathway and glyoxylate bypass[J]. Applied Microbiology and Biotechnology, 2018, 102(5):2179-2189.
[6]Alkim C, Trichez D, et al. The synthetic xylulose-1 phosphate pathway increases production of glycolic acid from xylose-rich sugar mixtures[J]. Biotechnology for Biofuels, 2016, 9(1):201.
[7]李宇展,刘伟,顾登平.草酸电还原研制乙醇酸[J].河北师范大学学报, 2003(4):385-387.
[8]Dahms AS. 3-Deoxy-D-pentulosonic acid aldolase and its role in a new pathway of D-xylose degradation[J]. Biochemical and biophysical research communications, 1974, 60(4):1433-1439.
[9]van Dijl JM, Hecker M. Bacillus subtilis:from soil bacterium to super-secreting cell factory[J]. Microb Cell Fact, 2013, 12:3.
[10]T?nnler S, Zamboni N, Kiraly C, et al. Screening of Bacillus subtilis transposon mutants with altered riboflavin production[J].Metabolic engineering, 2008, 10(5):216-226.
[11]李思杰,纪明华,赵利超,史吉平,孙俊松. N-乙酰神经氨酸合成途径在枯草芽孢杆菌的构建[J].食品工业科技, 2017,38(16):131-135.
[12]Liu H, Ramos KRM, Valdehuesa KNG, et al. Biosynthesis of ethylene glycol in Escherichia coli[J]. Applied Microbiology and Biotechnology, 2013, 97(8):3409-3417.
[13]Tai YS, Xiong M, Jambunathan P, et al. Engineering nonphosphorylative metabolism to generate lignocellulose-derived products[J]. Nature Chemical Biology, 2016, 12(4):247.
[14]Kolodkin-Gal I, Romero D, Cao S, et al. D-amino acids trigger biofilm disassembly[J]. Science, 2010, 328(5978):627-629.
[15]Cheng YL, Kalman LV, Kaiser D. The dsg gene of Myxococcus xanthus encodes a protein similar to translation initiation factor IF3[J]. Journal of Bacteriology, 1994, 176(5):1427-1433.
[16]Weimberg R. Pentose oxidation by Pseudomonas fragi[J]. J Biol Chem, 1961, 236(3):629-635.
[17]Meijnen JP, de Winde J H, et al. Establishment of oxidative D-xylose metabolism in Pseudomonas putida S12[J]. Applied and Environmental Microbiology, 2009, 75(9):2784-2791.