当前位置： SCI文献检索 > INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES期刊下所有文献 > Characterization of a recombinant d-allulose 3-epimerase from Agrobacterium sp. ATCC 31749 and identification of an important interfacial residue.

Characterization of a recombinant d-allulose 3-epimerase from Agrobacterium sp. ATCC 31749 and identification of an important interfacial residue.

Abstract：

:d-Allulose 3-epimerase (DAEase) catalyzes the epimerization between d-fructose and d-allulose. We had PCR-cloned and overexpressed the gene encoding Agrobacterium sp. ATCC 31749 DAEase (AsDAEase) in Escherichia coli. A high yield of active AsDAEase, 35,300U/L or 1350U/g of wet cells, was acquired with isopropyl β-d-1-thiogalactopyranoside induction at 20°C for 20h. Although only six residues including residue 234 located in tetrameric interface are different between AsDAEase and A. tumefaciens DAEase (AtDAEase), the specific activity of purified AsDAEase is much larger than that of AtDAEase. The optimal pHs and optimal temperatures of the purified recombinant AsDAEase are 7.5-8.0 and 55-60°C, respectively. The half-life of the enzyme is 267min at 55°C in the presence of 0.1mM Co2+, and the equilibrium ratio between d-allulose and d-fructose is 30:70 at 55°C. Besides characterizing AsDAEase, mutation N234D was constructed to assess its influence on activity. The specific activity of the purified N234D AsDAEase is only 25.5% of wild-type's activity, suggesting residue N234 is an important interfacial residue which substantially affects enzyme activity. The high specific activity and high expression yield of AsDAEase suggest its prospect to be applied in d-allulose production.

journal_name

Int J Biol Macromol

journal_title

International journal of biological macromolecules

authors

Tseng WC,Chen CN,Hsu CT,Lee HC,Fang HY,Wang MJ,Wu YH,Fang TY

doi

10.1016/j.ijbiomac.2018.02.036

subject

Has Abstract

pub_date

2018-06-01 00:00:00

pages

767-774

eissn

0141-8130

issn

1879-0003

pii

S0141-8130(18)30055-2

journal_volume

112

pub_type

杂志文章

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