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High-throughput Screening of Carbohydrate-degrading Enzymes Using Novel Insoluble Chromogenic Substrate Assay Kits
Published on: September 20, 2016
Screening for distinct xylan degrading enzymes in complex shake flask fermentation supernatants
M P Van Gool1, I Vancsó, H A Schols
1Wageningen University, Laboratory of Food Chemistry, Bomenweg 2, 6703 HD Wageningen, The Netherlands.
Bioresource Technology
|March 29, 2011
Summary
Efficiently degrading complex xylans requires multiple enzymes. A new screening method identifies specific xylan-modifying enzymes, revealing that no single fungus produces all necessary enzymes for complete xylan breakdown.
Area of Science:
- Biotechnology
- Enzymology
- Microbiology
Background:
- Complex xylan degradation necessitates a synergistic action of multiple enzymes.
- Traditional assays using reducing sugars or PNP substrates are insufficient for detecting enzymes crucial for complex xylan breakdown, including accessory enzymes.
Purpose of the Study:
- To develop and validate a novel screening method for identifying specific xylan-modifying enzymes in fungal isolates.
- To assess the enzymatic profiles of fungal soil isolates for their potential in complex xylan degradation.
Main Methods:
- Analysis of fermentation supernatants from 78 fungal soil isolates grown on wheat straw.
- Utilized High-Performance Liquid Chromatography (HPLC) and Mass Spectrometry (MS) for enzyme detection and characterization.
Main Results:
- The developed strategy effectively identified xylanases, arabinoxylan hydrolases, acetyl xylan esterases, and glucuronidases.
- No single fungal isolate possessed all enzymes required for complete substrate degradation.
- Some isolates showed high xylanase activity on linear xylan but lacked capability for complex xylan degradation, while others with lower xylanase levels secreted crucial accessory enzymes.
Conclusions:
- A new HPLC and MS-based screening method enables the detection of specific xylan-modifying enzymes.
- Fungal isolates exhibit diverse enzymatic capabilities, with no single strain capable of complete complex xylan degradation.
- The presence of accessory enzymes is critical for efficient complex xylan degradation, even in the presence of high levels of primary xylanases.
