Paper AS-MoA11
First use of ToF-SIMS for Screening Assays: Enzymes Active on Wood

Monday, October 31, 2011, 5:20 pm, Room 102

Proteomic and metagenomic studies are rapidly increasing the number of proteins available for enzymatic screening. However, current high-throughput enzyme assays have limited applicability for an important class of biochemical substrates – complex solid materials. The present work aims to utilize the strengths of Time-of-Flight Secondary Ion Mass Spectrometry for the direct measurement of enzyme activity on solid substrates. Particularly, ToF-SIMS is applied to the detection of wood-modifying enzymes.

Proof-of-principle ToF-SIMS enzyme assays were performed by immersing extracted wood fibers in solutions of commercial cellulase and laccase enzymes (utilizing water/buffer and denatured enzymes for controls). The laccase enzyme was also tested with and without several small molecule mediators. Principle Component Analysis (PCA) clearly distinguished cellulase tests from controls through the loss of polysaccharide peaks and relative enrichment of lignin peaks. Additionally, PCA distinguished laccase test samples (with mediator) from controls through a shift in lignin-characteristic peaks. The active laccase was indicated by a relative decrease in guaiacyl-lignin and syringyl-lignin peak intensities and increase in generic aromatic peaks, resulting from the cleavage of hydroxyl and methoxy groups from lignin benzoid units.

These proof-of-principle assays demonstrate that ToF-SIMS is capable of providing yes/no screening information for enzyme activity on complex solid substrates, such as wood.

[1] R. E. Goacher, D. Jeremic, E. R. Master. Analytical Chemistry 83(3), 2011, 804-812.