Vitamin B12 (cobalamin) has important physiological functions, which is widely used in pharmaceutical, food and other industries. Ensifer adhaerens is one of the commonly used strains for the production of vitamin B12. The processing time of atmospheric and room temperature plasma (ARTP) was 80 s, and the output power was 100 W. Through two rounds of ARTP mutagenesis, preliminary screening and re-screening, a genetic stability mutant strain 46H-1 with high vitamin B12 production was screened. By fermentation in shaking flask for 7.5 d, the titer of vitamin B12 reached 110.25 mg/L, which increased by 24.8% compared with E. adhaerens. The established high-throughput screening method could apply to harvest the excellent performance mutant strains quickly, thus reducing the cost of production.
XIAO Zhiqiang
,
ZHU Yongming
,
LI Jianghua
,
ZHOU Jingwen
. High-throughput screening of Ensifer adhaerens with improved vitamin B12 yield[J]. Food and Fermentation Industries, 2021
, 47(19)
: 7
-11
.
DOI: 10.13995/j.cnki.11-1802/ts.026462
[1] MINOT G R,MURPHY W P.Treatment of pernicious anemia by a special diet[J].Jama,1983,250:3 328-3 335.
[2] SMITH E L.Presence of cobalt in the anti-pernicious anaemia factor[J].Nature,1948,162(4 108):144.
[3] 马蕙, 王丽丽,张春晓,等.维生素B12的生物合成、发酵生产与应用[J].生物工程学报,2008,24(6):927-932.
MA H,WANG L L,ZHANG C X,et al.Biosynthesis,fermentation and application of vitamin B12-A review[J].Chinese Journal of Biotechnology,2008,24(6):927-932.
[4] WARREN M J,RAUX E,SCHUBERT H L,et al.The biosynthesis of adenosylcobalamin (vitamin B-12)[J].Natural Product Reports,2002,19(4):390-412.
[5] CHAN C W,MONDRAGÓN A.Crystal structure of an atypical cobalamin riboswitch reveals RNA structural adaptability as basis for promiscuous ligand binding[J].Nucleic Acids Research,2020,48(13):7 569-7 583.
[6] MARSH E N G,PATTERSON D P,LI L,et al.Adenosyl radical:Reagent and catalyst in enzyme reactions[J].Chembiochem,2010,11:604-621.
[7] XIA W,CHEN W,PENG W F,et al.Industrial vitamin B12 production by Pseudomonas denitrificans using maltose syrup and corn steep liquor as the cost-effective fermentation substrates[J].Bioprocess and Biosystems Engineering,2015,38(6):1 065-1 073.
[8] WANG P,ZHANG Z W,JIAO Y J,et al.Improved propionic acid and 5,6-dimethylbenzimidazole control strategy for vitamin B12 fermentation by Propionibacterium freudenreichii[J].Journal of Biotechnology,2015,193:123-129.
[9] THI VU H,ITOH H,ISHII S,et al.Identification and phylogenetic characterization of cobalamin biosynthetic genes of Ensifer adhaerens[J].Microbes and Environments,2013,28(1):153-155.
[10] RYCHEN G,AQUILINA G,AZIMONTI G,et al.Safety and efficacy of vitamin B12 (in the form of cyanocobalamin) produced by Ensifer spp.as a feed additive for all animal species based on a dossier submitted by VITAC EEIG[J].Efsa Journal,2018,16:e05336.
[11] FANG H,LI D,KANG J,et al.Metabolic engineering of Escherichia coli for de novo biosynthesis of vitamin B12[J].Nature Communications,2018,9(1):4 917.
[12] VITRESCHAK A G,RODIONOV D A,MIRONOV A A,et al.Regulation of the vitamin B12 metabolism and transport in bacteria by a conserved RNA structural element[J].RNA,2003,9(9):1 084-1 097.
[13] RATHORE D S,LOPEZ-VERNAZA M A,DOOHAN F,et al.Profiling antibiotic resistance and electrotransformation potential of Ensifer adhaerens OV14;a non-Agrobacterium species capable of efficient rates of plant transformation[J].FEMS Microbiology Letters,2015,362(17):fnv126.
[14] ZHANG X,ZHANG X F,LI H P,et al.Atmospheric and room temperature plasma (ARTP) as a new powerful mutagenesis tool[J].Applied Microbiology and Biotechnology,2014,98(12):5 387-5 396.
[15] ALVAREZ J C.High-throughput docking as a source of novel drug leads[J].Current Opinion in Chemical Biology,2004,8:365-370.
[16] LIANG Y,ZHU Y,FANG Q,et al.Advances in high-throughput,automated techniques and instrumention for protein crystallization screening[J].Chinese Journal of Chromatography,2016,34:1 137-1 144.
[17] DU G S,FANG Q,DEN TOONDER J M J.Microfluidics for cell-based high throughput screening platforms-A review[J].Analytica Chimica Acta,2016,903:36-50.
[18] LUO Z S,ZENG W Z,DU G C,et al.A high-throughput screening procedure for enhancing pyruvate production in Candida glabrata by random mutagenesis[J].Bioprocess abd Biosystems Engineering,2017,40(5):693-701.
[19] CAO X,LUO Z,ZENG W,et al.Enhanced avermectin production by Streptomyces avermitilis ATCC 31267 using high-throughput screening aided by fluorescence-activated cell sorting[J].Applied Microbiology Biotechnolgy,2018,102:703-712.
[20] CAI Y Y,XIA M M,DONG H N,et al.Engineering a vitamin B-12 high-throughput screening system by riboswitch sensor in Sinorhizobium meliloti[J].BMC Biotechnology,2018,18:27.
[21] ZENG W,GUO L,XU S,et al.High-throughput screening technology in industrial biotechnology[J].Trends in Biotechnology,2020,38:888-906.
