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Expression and Activity Analysis of Fructosyltransferase from Aspergillus oryzae

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Abstract

The fructosyltransferase gene was isolated and cloned from Aspergillus oryzae. The gene was 1368 bp, which encoded a protein of 455 amino acids. To analyze the activity of the expressed fructosyltransferase, the pET32a-fructosyltransferase recombined plasmid was transformed into Escherichia coli BL21. The fructosyltransferase gene was successfully expressed by Isopropyl-β-d-thiogalactoside (IPTG) induction. The molecular weight of the expression protein was about 45 kDa. The optimal conditions of protein expression were 25 °C, 0.1 mM IPTG, and 8 h of inducing time. The optimal concentration of urea dealing with inclusion body was 2.5 M. The expressed protein exhibited a strong fructosyl transfer activity. These results showed that the expressed fructosyltransferas owned transferase activity, and could catalyze the synthesis of sucrose-6-acetate.

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Abbreviations

A. oryzae :

Aspergillus oryzae

E. coli :

Escherichia coli

IPTG:

Isopropyl-β-d-thiogalactoside

HPLC:

High performance liquid chromatography

FOS:

Fructooligosaccrides

PDA:

Potato dextrose agar

LB:

Luria–Bertani

PCR:

Polymerase chain reaction

SDS-PAGE:

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis

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Acknowledegments

This work was supported by Basic Research Project of Science and Technology Department of Henan province (No. 132102210409), and National Natural Science Foundation of China (No. 31172175). We thank Dr. Ming Li (University of Virginia, USA) for his critical revising and suggestions.

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Authors and Affiliations

  1. College of Food and Bioengineering, Zhengzhou University of Light Industry, Number 5, Dongfeng Road, Zhengzhou, 450002, China

    Lihong Guan, Liping Chen, Nu Zhang & Yawei Han

  2. College of Life Science and Technology, Xinxiang Medical University, Xinxiang, China

    Lihong Guan

  3. Jilin Tobacco Industrial Co. Ltd., Yanji, China

    Yongsen Chen

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  1. Lihong Guan
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  2. Liping Chen
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Correspondence to Yawei Han.

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Guan, L., Chen, L., Chen, Y. et al. Expression and Activity Analysis of Fructosyltransferase from Aspergillus oryzae . Protein J 36, 352–360 (2017). https://doi.org/10.1007/s10930-017-9725-y

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  • DOI: https://doi.org/10.1007/s10930-017-9725-y

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