Cloning and expression of an endoglucanase gene from the thermotolerant fungus Aspergillus fumigatus DBiNU-1 in Kluyveromyces lactis

Abstract

An intronless endoglucanase from thermotolerant Aspergillus fumigatus DBINU-1 was cloned, characterized and expressed in the yeast Kluyveromyces lactis. The full-length open reading frame of the endoglucanase gene from A. fumigatus DBiNU-1, designated Cel7, was 1383 nucleotides in length and encoded a protein of 460 amino acid residues. The predicted molecular weight and the isoelectric point of the A. fumigatus Cel7 gene product were 48.19 kDa and 5.03, respectively. A catalytic domain in the N-terminal region and a fungal type cellulose-binding domain/module in the C-terminal region were detected in the predicted polypeptide sequences. Furthermore, a signal peptide with 20 amino acid residues at the N-terminus was also detected in the deduced amino acid sequences of the endoglucanase from A. fumigatus DBiNU-1. The endoglucanase from A. fumigatus DBiNU-1 was successfully expressed in K. lactis, and the purified recombinant enzyme exhibited its maximum activity at pH 5.0 and 60 °C. The enzyme was very stable in a pH range from 4.0 to 8.0 and a temperature range from 30 to 60 °C. These features make it suitable for application in the paper, biofuel, and other chemical production industries that use cellulosic materials.

Trade-Offs Between Growth Rate and Other Fungal Traits

“…Invertase 32 Breakdown of sucrose (Boddy et al, 1993) α-Glucosidase α-Glucosidase 15; α-Glucosidase 31 Breakdown of starch (Nakamura et al, 1997) Amylase Amylase 88 Breakdown of starch (Korman et al, 1990) β-Xylosidase β-Xylosidase 43 Breakdown of hemicellulose (Semenova et al, 2009) β-Glucosidases β-Glucosidase 1 Breakdown of cellulose (Znameroski et al, 2012) Cellobiohydrolases Cellobiohydrolase 6; Cellobiohydrolase 7 Breakdown of cellulose (Ilmén et al, 1997) Endoglucanases Endoglucanase 9; Endoglucanase 12 Breakdown of cellulose (Rungrattanakasin et al, 2018) α-Mannosidase Glycoprotein synthesis 92; α-mannanase 76…”

Thermostable Cellulases / Xylanases From Thermophilic and Hyperthermophilic Microorganisms: Current Perspective

“…K. lactis has so many advantages as a platform for the expression of a recombinant protein, which include easy genetic manipulation, application of both integrative and episomal expression vectors, it is known to produce a protein that is very useful in the food and dairy industry ( Spohner et al, 2016 ). A thermostable endoglucanase was clone from Aspergillus fumigatus DBINU-1 and it was heterologously express in K. lactis this recombinant enzyme has optimal activity at pH 5.0 and 60°C ( Rungrattanakasin et al, 2018 ). Similarly, an alkali-thermostable xylanase was cloned from Bacillus pumilus and it was expressed in K. lactis this shows very good stability at pH 12.0 retaining 74% of it Activity after incubation for 2 h it is suitable for application in pulp and paper industry ( Thomas et al, 2014b ).…”

Recent Developments in Industrial Mycozymes: A Current Appraisal

“… (Karima et al 2019 ) Endoglucanase Aspergillus fumigatus Cel7 pKLAC2 PLAC4-PBI K. lactis GG799 Application in the paper industry and in saccharification processes for the production of biofuel and other chemicals using cellulosic materials. (Rungrattanakasina et al 2018 ) Cellobiohydrolase Trichoderma virens cbhI ANIp5MS Gla Pr A. niger strain PY11 Useful in complex biomass degradation. (Fatimi et al 2018 ) Xylanase Aspergillus oryzae XynF1 pET28a T7 E. coli BL21 (DE3) Pulp biobleaching in paper industry (Bhardwaj et al 2020 ) Mannanase Aspergillus kawachii ManAK pPICZαA AOX1 P. pastoris X33 Mannooligosaccharide (MOS) preparation and application in food and feed area.…”