Premium
A Novel Thermotolerant β‐glucosidase from Aspergillus nidulans Has Activity Across a Broad pH Profile and a Likely Bacterial Origin
Author(s) -
Auta Richard,
Radecka Iza,
Hooley Paul
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.1070.1
Subject(s) - chemistry , aspergillus nidulans , hydrolysis , thermostability , ammonium , isoelectric point , enzyme , sephadex , chromatography , size exclusion chromatography , biochemistry , gene , mutant , organic chemistry
This study aims at cloning, expression and biochemical characterization of recombinant β‐glucosidases from Aspergillus nidulans AN1804 gene for biofuel production. Purification was conducted using ammonium sulphate precipitation and anion exchange chromatography on a DEAE‐Sephadex A‐50 column. The enzyme was purified 2.25 fold from the crude extract with a 33.33% recovery yield. The protein migrated homogeneously as a single band on SDS‐PAGE stained with Sterling rapid silver stain and the molecular weight was determined to be approximately 100 kDa. It was optimally active at 50 °C and pH 5.5, though it had a broad pH range of pH 3.0 – 10.0. The presence of CoCl 2 , CaCl 2 , FeCl 3 , FeCl 2 and ZnCl 2 slightly enhanced the activity of β‐glucosidase but was slightly decreased in the presence of HgCl 2 . β‐glucosidase showed very high affinity to para‐Nitrophenyl β‐D‐glucopyranoside (pNPG). K m and V max for the hydrolysis of pNPG by β‐glucosidase was calculated as 0.59 mM and 2.04 μmole/ml/min respectively. An initial bioinformatics studies shows a rare occurrence of introns in AN1804, providing additional support to the argument of horizontal transfer from a bacterial source. The spectacular thermo‐tolerance and very broad pH profile of AN1804 β‐glucosidase could be the reason why the fungus has kept the bacterial gene. Support or Funding Information Personal funding