Gene overexpression and biochemical characterization of the biotechnologically relevant chlorogenic acid hydrolase from Aspergillus niger - Archive ouverte HAL Access content directly
Journal Articles Applied and Environmental Microbiology Year : 2007

Gene overexpression and biochemical characterization of the biotechnologically relevant chlorogenic acid hydrolase from Aspergillus niger

Abstract

The full-length gene encoding the chlorogenic acid hydrolase from Aspergillus niger CIRM BRFM 131 was cloned by PCR based on the genome of the strain A. niger CBS 513.88. The complete gene consists of 1715 bp and codes for a deduced protein of 512 amino acids with a molecular weight of 55264 Da and an acidic pI of 4.6. The gene was successfully cloned and overexpressed in A. niger to yield 1.25 g L-1, i.e. 330-fold higher than compared to the production of the wild-type strain A. niger CIRM BRFM131. The histidine-tagged recombinant ChloE was purified to homogeneity via a single chromatography step and characterized for the main biochemical properties. The molecular size of the protein checked by mass spectroscopy was 74553 Da, suggesting the presence of glycosylation. ChloE is assembled in a tetrameric form with several acidic isoforms with pI around 4.55 and 5.2. Other characteristics such as optimal pH and temperature were found to be similar to those determined for the previously characterized chlorogenic acid hydrolase of A. niger CIRM BRFM 131. However, there was a significant temperature stability difference in favor of the recombinant protein. ChloE exhibits a catalytic efficiency of 12.5 x 106 M-1 s-1 towards chlorogenic acid (CGA) and its ability to release caffeic acid from CGA present in agricultural by-products such as apple marc and coffee pulp was clearly demonstrated, confirming the high potential of this enzyme.

Dates and versions

hal-02658406 , version 1 (30-05-2020)

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Isabelle Benoit, Michèle Asther, Yves Bourne, David Navarro, Stéphane Canaan, et al.. Gene overexpression and biochemical characterization of the biotechnologically relevant chlorogenic acid hydrolase from Aspergillus niger. Applied and Environmental Microbiology, 2007, 73 (17), pp.5624-5632. ⟨10.1128/AEM.00374-07⟩. ⟨hal-02658406⟩
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