Please wait a minute...

中国生物工程杂志

China Biotechnology
China Biotechnology  2016, Vol. 36 Issue (1): 47-54    DOI: 10.13523/j.cb.20160107
    
Effects of D194G Mutant on meso-2, 3-Butanediol Dehydrogenase Catalytic Properties
HAO Wen-bo1,2, JI Fang-ling1, WANG Jing-yun1, ZHANG Yue1, WANG Tian-qi1, CHE Wen-shi2, BAO Yong-ming1
1. School of Life Science and Biotechnology, Dalian University of Technology, Dalian 116024, China;
2. Department of Physics and Chemistry, Heihe University, Heihe 164300, China
Download: HTML   PDF(1433KB) HTML
Export: BibTeX | EndNote (RIS)      

Abstract  

Objective: To compare the activity and kinetic parameters of meso-2,3-Butanediol dehydrogenase (BDH) from Enterobacter aerogenes (E. a-BDH) and Bacillus subtilis (B. s-BDH), and analysis the influences of the residue D194 on catalytic properties of BDH. Methods: E. a-BDH and D194G B. s-BDH were expressed in E. coli BL21 (DE3), and purified by HiTrap Q FF anion-exchange and Superdex 75 gel column. MALDI-TOF MS was used to determine the molecular weight. The enzyme activity, coenzyme and substrate specificity, optimum pH, temperature, and kinetic parameters of BDH were investigated by monitoring changes in absorbance of NADH/NAD+ redox reaction. Results: The recombinant E. a-BDH and D194G B. s-BDH are homo-tetramer. Their nucleotide sequences exhibit two different bases (g.27A/T and g.581A/G), and g.581A/G results in an amino acid change (p.D194G). D194G B. s-BDH activity is about 2.3% of E. a-BDH, and lost the ability of oxidation of meso-2, 3-butanediol. Acetoin/NADH is the optimal substrate of BDH, but Km of D194G B. s-BDH is 5.63 times greater than that of E. a-BDH. Conclusion: D194G mutation reduces the BDH activity.



Key wordsmeso-2 3-butanediol dehydrogenase      Acetoin      Bacillus subtilis     
Received: 04 August 2015      Published: 11 January 2016
ZTFLH:  Q78  
Cite this article:

HAO Wen-bo, JI Fang-ling, WANG Jing-yun, ZHANG Yue, WANG Tian-qi, CHE Wen-shi, BAO Yong-ming. Effects of D194G Mutant on meso-2, 3-Butanediol Dehydrogenase Catalytic Properties. China Biotechnology, 2016, 36(1): 47-54.

URL:

https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20160107     OR     https://manu60.magtech.com.cn/biotech/Y2016/V36/I1/47

[1] Xiao Z J, Lu J R. Strategies for enhancing fermentative production of acetoin: A review. Biotechnology Advances, 2014, 32(2): 492-503.
[2] Sun J, Zhang L, Rao B, et al. Enhanced acetoin production by Serratia marcescens H32 with expression of a water-forming NADH oxidase. Bioresource Technology, 2012, 119(7): 94-98.
[3] Liu Y, Zhang S, Yong Y, et al. Efficient production of acetoin by the newly isolated Bacillus licheniformis strain MEL09. Process Biochemistry, 2011, 46(1): 390-394.
[4] Teixeira R M, Cavalheiro D, Ninow J L, et al. Optimization of acetoin production by Hanseniaspora guilliermondii using experimental design. Brazilian Journal of Chemical Engineering, 2002, 19(2): 181-186.
[5] Zhang Y J, Li S B, Liu L M, et al. Acetoin production enhanced by manipulating carbon flux in a newly isolated Bacillus amyloliquefaciens. Bioresource Technology, 2013, 130(1): 256-260.
[6] Luo Q L, Wu J, Wu M C. Enhanced acetoin production by Bacillus amyloliquefaciens through improved acetoin tolerance. Process Biochemistry, 2014, 49(8): 1223-1230.
[7] Zhang X, Zhang R Z, Bao T, et al. The rebalanced pathway significantly enhances acetoin production by disruption of acetoin reductase gene and moderate-expression of a new water-forming NADH oxidase in Bacillus subtilis. Merabolic Engineering, 2014, 23(2): 34-41.
[8] Li S B, Xu N, Liu L M, et al. Engineering of carboligase activity reaction in Candida glabrata for acetoin production. Merabolic Engineering, 2014, 22(3): 32-39.
[9] Borim K, Lee S, Park J, et al. Enhanced 2, 3-butanediol production in recombinant Klebsiella pneumoniae via overexpression of synthesis-related genes. Journal of Microbiology and Biotechnology, 2012, 22(9): 1258-1263.
[10] Han S H, Lee J E, Park K, et al. Production of 2, 3-butanediol by a low-acid producing Klebsiella oxytoca NBRF4. New Biotechnology, 2013, 30(2): 166-172.
[11] Yang T W, Rao Z M, Zhang X, et al. Effects of corn steep liquor on production of 2, 3-butanediol and acetoin by Bacillus subtilis. Process Biochemistry, 2013, 48(11): 1610-1617.
[12] Nakashima N, Akita H, Hoshino T. Establishment of a novel gene expression method, BICES (biomass-inducible chromosome-based expression system), and its application to the production of 2, 3-butanediol and acetoin, Merabolic Engineering, 2014, 25(1): 204-214.
[13] Ji X J, Huang H, Ouyang P K. Microbial 2, 3-butanediol production: a state-of-the-art review. Biotechnology Advances, 2011, 29(3): 351-364.
[14] Celinska E, Grajek W. Biotechnological production of 2, 3-butanediol - current state and prospects, Biotechnology Advances, 2009, 27(6): 715-725.
[15] Li L X, Wang Y, Zhang L J, et al. Biocatalytic production of (2S, 3S)-2, 3-butanediol from diacetyl using whole cells of engineered Escherichia coli. Bioresource Technology, 2011, 115(4): 111-116.
[16] Zhang LY, Yang Y L, Sun J A, et al. Microbial production of 2, 3-butanediol by a mutagenized strain of Serratia marcescens H30. Bioresource Technology, 2010, 101(6): 1961-1967.
[17] Yu B, Sun J B, Bommareddy R R, et al. Novel (2R, 3R)-2, 3-butanediol dehydrogenase from potential industrial strain Paenibacillus polymyxa ATCC 12321. Applied and Environmental Microbiology, 2011, 77(12): 4230-4233.
[18] González E, Fernández M R, Larroy C, et al. Characterization of a (2R, 3R)-2,3-butanediol dehydrogenase as the Saccharomyces cerevisiae YAL060W gene product. Journal of Biological Chemistry, 2000, 275(46): 35876-35885.
[19] Hao W B, Ji F L, Wang J Y, et al. Biochemical characterization of unusual meso-2, 3-butanediol dehydrogenase from a strain of Bacillus subtilis. Journal of Molecular Catalysis B-Enzymatic, 2014, 109(1): 184-190.
[20] Otagiri M, Kurisu G, Ui S, et al. Crystal structure of meso-2, 3-butanediol dehydrogenase in a complex with NAD+ and inhibitor mercaptoethanol at 1.7 A resolution for understanding of chiral substrate recognition mechanisms. Journal of Biochemistry, 2001, 129(2): 205-208.
[21] Giovannini P P, Medici A, Bergamini C M, et al. Properties of diacetyl (acetoin) reductase from Bacillus stearothermophilus. Bioorganic & Medicinal Chemistry Letters, 1996, 4(8): 1197-1201.
[22] Schwarz J G, Hang Y D. Purification and characterization of diacetyl reductase from Kluyveromyces marxianus. Letters in Applied Microbiology, 1994, 18(5): 272-276.
[23] Ui S, OkajimaY, Mimura A, et al. Sequence analysis of the gene for and characterization of D-acetoin forming meso-2,3-butanediol dehydrogenase of Klebsiella pneumoniae expressed in Escherichia coli. Journal of Fermentation and Bioengineering, 1997, 83(97): 32-37.
[24] Filling C, Berndt K D, Benach J, et al. Critical residues for structure and catalysis in short-chain dehydrogenases/reductases. Journal of Biological Chemistry, 2002, 277(28): 25677-25684.
[25] Ghosh D, Weeks C M, Grochulski P, et al. Three-dimensional structure of holo 3 alpha, 20 beta-hydroxsteroid dehydrogenase: a member of short-chain dehydrogenase family. Proceedings of the National Academy of Sciences, 1991, 88(22): 10064-10068.
[26] Tanaka N, Nonaka T, Nakanishi M, et al. Crystal structure of the ternary complex of mouse lung carbonyl reductase at 1.8 ? resolution: the structural origin of coenzyme specificity in the short-chain dehydrogenase/reductase family. Structure, 1996, 4(1): 33-45.
[27] Otagiri M, Ui S, Takusagawa Y, et al. Structural basis for chiral substrate recognition by two 2,3-butanediol dehydrogenases. FEBS Letters, 2010, 584(1): 219-223.
[28] Rao B, Zhang L Y, Sun J A, et al. Characterization and regulation of the 2, 3-butanediol pathway in Serratia marcescens. Applied Microbiology and Biotechnology, 2012, 93(5): 2147-2159.

[1] Jian-xiu LI,Xian-rui CHEN,Xiao-ling CHEN,Yan-yan HUANG,Qi-wen MO,Neng-zhong XIE,Ri-bo HUANG. Construct Whole-cell Biocatalyst and Produce (S)-Acetoin via Synthetic Biology Strategy[J]. China Biotechnology, 2019, 39(4): 60-68.
[2] ZHAO Yi-jin, WANG Teng-fei, WANG Jun-qing, WANG Rui-ming. Surface Display of Tres Using CotC as a Molecular Vector on Bacillus subtilis Spores[J]. China Biotechnology, 2017, 37(1): 71-80.
[3] HU Gui-yuan, YANG Tao-wei, RAO Zhi-ming, LIU Mei, XU Mei-juan, ZHANG Xian. Improved Production of 2,3-Butanediol by Enhancing the Level of Intracellular NADH and Activity of Acetoin Reductase[J]. China Biotechnology, 2016, 36(6): 57-64.
[4] LIU Xiao-fei, FU Jing, HUO Guang-xin, ZHANG Bo, WANG Zhi-wen, CHEN Tao. Latest Advances of Microbial Production of Platform Chemical Acetoin[J]. China Biotechnology, 2015, 35(10): 91-99.
[5] XIE Zhi-dan, FAN Wen, JIA Dong-cheng, YANG Na, XIA Zheng-yuan, QIAO Min. Recent Developments in Spore Surface Display of Bacillus subtilis[J]. China Biotechnology, 2014, 34(8): 105-111.
[6] LIU Hui-li, LI Yuan-yuan, JU Rui-cheng, ZHAO Hong-tao, YANG Qing. Isolation, Identification and Fermentation Optimization of Antagonistic Bacillus subtilis KC-5[J]. China Biotechnology, 2014, 34(3): 96-102.
[7] HUANG Xiang-feng, ZHAN Peng-ju, PENG Kai-ming, LIU Jia, LU Li-jun. Study on the Influence of Iron Dosage in the Medium on Fermentation of Lipopeptide Produced by Bacillus subtilis CICC 23659[J]. China Biotechnology, 2013, 33(6): 52-61.
[8] WU Hai-li, ZHANG San-jun, DU Bing, QIAN Min, REN Hua. Expression, Purification and Biological Activity of Arginine Mutants of Bacillus subtilis RecQ Helicase[J]. China Biotechnology, 2013, 33(12): 29-34.
[9] LIU Jin-xia, Ll Na, DU Wen-jing, WU Jian-rong, LI Jin, DING Pin, SHEN Si-yuan, ZHANG Jian-jun. The Research of the Highly Effective Biocontrol Emulsion with Bacillus subtilis[J]. China Biotechnology, 2011, 31(9): 69-75.
[10] ZHANG Yang, DU Shan-shan, XIE Xi-xan, XU Qing-yang, CHEN Ning. Effect of purA Gene Overexpression on Adenosine Accumulation[J]. China Biotechnology, 2011, 31(12): 22-26.
[11] LI Xiao-jing, DUAN Yun-xia. Application of Metabolic Engineering in Riboflavin Production[J]. China Biotechnology, 2011, 31(02): 130-138.