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A screening approach for assessing lytic polysaccharide monooxygenase activity in fungal strains

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journal contribution
posted on 2019-01-01, 00:00 authored by P Dixit, B Basu, Munish PuriMunish Puri, D K Tuli, A S Mathur, Colin BarrowColin Barrow
Background: Efforts to develop efficient lignocellulose-degrading enzymatic preparations have led to the relatively recent discovery of a new class of novel cellulase boosters, termed lytic polysaccharide monoxygenases (LPMOs). These enzymes are copper-dependent metalloenzymes that initiate the biomass deconstruction process and subsequently work together with cellulases, hemicellulases, and other accessory enzymes to enhance their hydrolytic action. Given their wide distribution and diversity, screening and isolation of potent LPMOs from natural fungal diversity may provide an important avenue for increasing the efficiency of cellulases and thereby decreasing cellulosic ethanol production costs. However, methods for quick screening and detection are still not widely available. In this article, a simple and sensitive method is described by combining nonhydrolytic activity enhancement followed by LC-MS-based quantitation of LPMOs. Results: In this study, a screening approach has been developed for the detection of nonhydrolytic cellulase-enhancing enzymes in crude fungal supernatants. With the application of a saturating benchmark cocktail of Celluclast 1.5L, fungal isolates were selected which had the capability of hydrolyzing pretreated rice straw by their synergistic enzyme fractions. Subsequently, these fungal extracts along with an LPMO-enriched commercial enzyme were investigated for their ability to produce Type I LPMO activity. An LC-MS-based methodology was applied to quantitate gluconic acid in enzymatic hydrolysates as an indirect measurement of Type I LPMO activity. Conclusion: The present study describes an LC-MS-based separation method to detect and quantitate LPMO activity in a commercial enzyme. This method was also applied to screen fungal extracts. The developed screening strategy has enabled detection of LPMO activity in two industrially important Penicillium strains.

History

Journal

Biotechnology for biofuels

Volume

12

Article number

185

Pagination

1 - 16

Publisher

BioMed Central

Location

London, Eng.

ISSN

1754-6834

Language

eng

Publication classification

C1 Refereed article in a scholarly journal

Copyright notice

2019, The Author(s)