Extracellular aldonic acid lactonase PoALAC and application thereof

A technology of lactonase and aldonic acid, applied in the fields of application, hydrolase, sugar production, etc.

Active Publication Date: 2017-03-22
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] After searching, there is no report about the e

Method used

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  • Extracellular aldonic acid lactonase PoALAC and application thereof
  • Extracellular aldonic acid lactonase PoALAC and application thereof
  • Extracellular aldonic acid lactonase PoALAC and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1: Preparation of extracellular aldonic lactonase PoALAC

[0030] 1. Microbial source and carrier

[0031] The original starting fungus Penicillium oxalicum (Penicillium oxalicum) wild strain 114-2 is a strain deposited by the applicant during the previous patent application. This strain was deposited in the General Microbiology Center (CGMCC) of the China Microbial Strain Collection Management Committee on September 28, 2011, and its deposit number is CGMCC No. 5302.

[0032] The Pichia expression vector pPIC9K and Pichia strain GS115 were purchased from Invitrogen.

[0033] 2. Medium

[0034] Mycelial growth medium: glucose 20.0g / L, KH 2 PO 4 3.0g / L, (NH 4 ) 2 SO 4 2.0g / L, MgSO 4 ﹒ 7H 2 O0.5g / L, CaCl 2 0.5g / L, FeSO 4 ﹒ 7H 2 O 0.0075g / L, MnSO 4 ﹒ H 2 O 0.0025g / L, ZnSO 4 ﹒ 7H 2 O0.0036g / L, CoCl 2 ﹒ 6H 2 O 0.0037g / L.

[0035] Induction medium: Replace the 20.0g / L glucose in the mycelial growth medium with 1% cellulose and 1% glucose.

[0036] E. coli culture medium ...

Embodiment 2

[0060] Example 2: Application of extracellular aldonic lactonase PoALAC in synergistic degradation of cellulose.

[0061] 1. Synergistic experiment of aldonic acid lactonase PoALAC and polysaccharide monooxygenase PoLPMO1 and β-glucosidase PoBGL1

[0062] The preparation method of phosphoric acid swollen cellulose: Add 2g of microcrystalline cellulose and 6mL of double distilled water into a 50mL centrifuge tube, stir thoroughly to make the cellulose powder into a uniform slurry. Add 10 mL of ice-cold 86% phosphoric acid and stir thoroughly to fully dissolve the cellulose and the mixture becomes clear. Place it in ice for 1 hour with constant stirring. Finally, add ice water and stir vigorously until the final concentration of phosphoric acid reaches 83.2% (wt / vol). At this time, a large amount of white gelatinous substance precipitates out. Centrifuge at 5000rpm and 4℃ for 20min. Wash the jelly with ice-cold double distilled water 3 times, and finally use 2M Na 2 CO 3 Neutralize ...

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Abstract

The invention discloses an extracellular aldonic acid lactonase PoALAC. An amino acid sequence thereof is as shown in SEQ ID NO.1 and a nucleotide sequence thereof is as shown in SEQ ID NO.2. The invention also discloses an application of the extracellular aldonic acid lactonase PoALAC in synergic degradation of celluloses. The specific enzyme activity can reach up to 173009IU/mg. After the extracellular aldonic acid lactonase PoALAC is added into a cellulose system, the extracellular aldonic acid lactonase PoALAC has a synergic effect with polysaccharide monooxygenase and beta-glucosidase and the degrading effects for different pre-treated natural lignocellulose substrates are promoted, so that the extracellular aldonic acid lactonase PoALAC has huge application potential and economic benefit in the industries of biological energy source and bio-based chemicals.

Description

Technical field [0001] The invention relates to an extracellular aldonic lactonase, in particular to an extracellular aldonic lactonase PoALAC and its application, and belongs to the field of genetic engineering. Background technique [0002] Lignocellulosic biomass is the most abundant renewable resource in the world and can be used to produce bioethanol fuel and other value-added products. The degradation of cellulose requires the synergistic action of endoglucanase cellobiohydrolase to convert it into cellobiose, which is then hydrolyzed into glucose by β-glucosidase. In recent years, it has been discovered that a lysing polysaccharide monooxygenase (LPMOs) can break the cellulose chain through oxidation, thereby synergistically degrading lignocellulose with other cellulolytic enzymes. Studies have shown that the hydrolysates of polysaccharide monooxygenase include cellooligosaccharides, aldonic acid lactones and aldonic acid. As we all know, aldonic acid lactone has inhibit...

Claims

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Application Information

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IPC IPC(8): C12N9/18C12N15/55C12N15/81C12N1/19C12P19/14C12P19/02C12R1/84
CPCC12N9/18C12P19/02C12P19/14C13K1/02
Inventor 曲音波彭圣娟刘国栋李雪芝
Owner SHANDONG UNIV
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