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Volume 69, Issue 8


Isolation of cDNA and upstream sequence of a gene encoding trehalose-6-phosphate synthase 1 from Beauveria bassiana and its functional identification in Pichia pastoris

Ling Xie
  • Anhui Provincial Key Laboratory of Microbial Pest Control, Anhui Agricultural University, Hefei, 230036, People’s Republic of China
  • School of Life Sciences, Anqing Teachers College, Anqing, 246011, People’s Republic of China
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/ Hongmei Chen / Zhangxun Wang
  • Anhui Provincial Key Laboratory of Microbial Pest Control, Anhui Agricultural University, Hefei, 230036, People’s Republic of China
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/ Bo Huang
  • Anhui Provincial Key Laboratory of Microbial Pest Control, Anhui Agricultural University, Hefei, 230036, People’s Republic of China
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Published Online: 2014-08-19 | DOI: https://doi.org/10.2478/s11756-014-0407-3


Trehalose is an important molecule in fungal cells that helps to protect against various environmental stresses. In most fungi, trehalose-6-phosphate synthase 1 (TPS1) catalyzes the synthesis of trehalose-6-phosphate, and is the key enzyme for biosynthesis of this sugar. In this study, the full-length Beauveria bassiana tps1 gene sequence was determined. Full-length Bbtps1 (1,906 bp) included a 1,563 bp open reading frame that contained a 55 bp intron located between deduced amino acids 104 and 105. Bioinformatics analysis predicted that the BbTPS1 protein comprised 520 residues with a calculated pI value of 5.63 and a molecular weight of 58.3 kDa. Using DNA walking experiments, we determined the 2,963 bp upstream sequence that included several typical promoter elements and putative transcription factor binding sites, such as TATA-box, GC-box, Oct-1, CRE-BP, CdxA, and GATA. Stress-response and heat-shock elements were also found in this upstream sequence. Recombinant BbTPS1 was expressed in Pichia pastoris GS115 in order to probe the function of Bbtps1. SDS-PAGE analysis showed that the expressed protein had a molecular weight of approximately 60 kDa as expected. Enzymatic activity measurements revealed specific TPS1 activity that peaked at 1.38 U/mL at 96 h. This work provides a basis for further functional investigation of the mechanism of trehalose anabolism in B. bassiana. It could also assist the construction of engineered B. bassiana strains with enhanced stress tolerance.

Keywords: Beauveria bassiana; trehalose-6-phosphate synthase 1; gene cloning; upstream sequence; Pichia pastoris

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About the article

Published Online: 2014-08-19

Published in Print: 2014-08-01

Citation Information: Biologia, Volume 69, Issue 8, Pages 959–967, ISSN (Online) 1336-9563, DOI: https://doi.org/10.2478/s11756-014-0407-3.

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