重组木聚糖酶的高密度发酵及其定向制备低聚木糖

doi:10.3969/j.issn.0253-2417.2024.01.013

Abstract

Abstract:

In order to improve the enzymatic hydrolysis efficiency of poplar xylan to xylo-oligosaccharides and reduce the cost of enzymatic preparation of xylo-oligosaccharides, this study focused on the induced expression conditions of Bacteroides-derived xylanase PdoXyn10A produced by high-density fermentation of recombinant Escherichia coli (PET-PdoXyn10A-DE3) and the process conditions of enzymatic hydrolysis to prepare xylo-oligosaccharides. The effects of the inducer concentration, the induced optical density (OD₆₀₀) value, induction temperature and induction pH value on the production of recombinant xylanase PdoXyn10A by PET-PdoXyn10A-DE3 were studied under 5 L fermenter level. The effects of reaction temperature and pH value on the enzyme activity of corncob xylanase during hydrolysis and the effects of enzyme addition and hydrolysis time on the preparation of xylo-oligosaccharide from poplar acetic acid hydrolysate were determined. The results showed that the activity of recombinase reached 362.67 U/mL and the biomass of recombinant Escherichia coli reached 28.83 g/L under high density fermentation at the optimal induction conditions with 0.25 mmol/L of inducer concentration, induction OD₆₀₀ value of 55, induction temperature 34 ℃ and induction pH value 7.0. The optimum conditions for the preparation of xylooligosaccharides by enzymatic hydrolysis were as follows: enzyme reaction temperature of 40 ℃, pH value of 5.2, the enzyme dosage of 12 U/mL, and reaction time of 6 h. Under these conditions, the content of xylo-oligosaccharides was 3.21 g/L, the content of xylobiose, xylotriose, xylotetraose, xylopentaose, xylohexaose were 1.09, 0.97, 0.84, 0.18, 0.13 g/L, respectively, and the content of xylose was 1.82 g/L.

Key words: recombinant xylanase, high-density fermentation, acetic acid hydrothermal reaction, xylooligosaccharide, enzyme hydrolysis condition

CLC Number:

TQ35

Jian ZHAO, Jingcong XIE, Yue JIANG, Jing YANG, Hao XU, Ning ZHANG. High Density Fermentation of Recombinant Xylanase and Its Directional Preparation of Xylooligosaccharides[J]. Chemistry and Industry of Forest Products, 2024, 44(1): 97-104.

Figures/Tables 4

Table 1

Fig.1

Table 2

Table 3

References 30

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诱导表达条件 induced expression condition		菌体生物量/(g·L^-1) bacterial biomass	酶活力/(U·mL^-1) enzyme activity
诱导剂浓度 IPTG concentration	0.10 mmol/L	24.42±0.03	218.11±3.16
	0.25 mmol/L	22.95±0.04	277.37±5.77
	0.50 mmol/L	21.96±0.02	263.51±6.08
	0.75 mmol/L	21.15±0.03	249.34±5.76
诱导OD₆₀₀ induction OD₆₀₀	15	20.32±0.77	210.15±2.14
	35	22.95±0.41	277.37±5.77
	55	23.88±0.23	317.38±5.09
	75	24.27±0.27	289.34±3.66
诱导温度 induction temperature	28 ℃	23.88±0.20	317.38±5.09
	31 ℃	25.17±0.43	323.34±3.25
	34 ℃	25.83±0.25	362.67±3.51
	37 ℃	31.53±0.47	308.05±4.89
诱导pH值 induction pH value	6.2	22.56±0.36	282.23±6.22
	6.6	27.21±0.41	318.42±5.42
	7.0	25.83±0.25	362.67±3.51
	7.4	28.08±0.37	332.91±7.08

酶添加量/(U·mL^-1) enzyme dosage	木糖/(g·L^-1) xylose	木二糖/(g·L^-1) xylobiose	木三糖/(g·L^-1) xylotriose	木四糖/(g·L^-1) xylotetraose	木五糖/(g·L^-1) xylopentaose	木六糖/(g·L^-1) xylohexaose	低聚木糖/(g·L^-1) XOS
0	1.62	0.64	0.62	0.60	0.34	0.32	2.52
4	1.73	0.75	0.74	0.62	0.35	0.34	2.80
8	1.76	0.89	0.87	0.60	0.33	0.26	2.95
12	1.80	1.03	0.96	0.82	0.21	0.18	3.20
16	2.13	1.00	0.93	0.84	0.11	0.09	2.97

水解反应时间/h reaction time	木糖/(g·L^-1) xylose	木二糖/(g·L^-1) xylobiose	木三糖/(g·L^-1) xylotriose	木四糖/(g·L^-1) xylotetraose	木五糖/(g·L^-1) xylopentaose	木六糖/(g·L^-1) xylohexaose	低聚木糖/(g·L^-1) XOS
0	1.62	0.64	0.62	0.60	0.34	0.32	2.52
4	1.74	0.87	0.85	0.63	0.34	0.32	3.01
5	1.80	1.03	0.96	0.82	0.21	0.17	3.19
6	1.82	1.09	0.97	0.84	0.18	0.13	3.21
7	1.91	1.10	0.95	0.86	0.16	0.12	3.19

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High Density Fermentation of Recombinant Xylanase and Its Directional Preparation of Xylooligosaccharides

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