SBIR Phase I: A Novel Probe for Biofilms

The broader impact of this Small Business Innovation Research (SBIR) Phase I project will be to improve performance of pulp and paper mills, especially small mills in rural areas, and to improve the quality of their products. The main components of wood are cellulose (used for paper), hemicellulose, and lignin (which makes paper turn yellow and brittle with age). Removal of lignin is key to papermaking, but it is typically attached to the hemicellulose. The project advances a technology to measure how much lignin is left in wood pulp. In the longer term, the probe may also detect biofilms which are layers of carbohydrates and other materials in which bacteria and fungi can grow. Water treatment facilities and processors of plant-derived materials routinely find that microorganisms naturally present in their starting materials form biofilms that can accumulate on machinery or contaminate the products. This technology has wide-ranging applications. The proposed project creates a peptide probe for non-glycosidic bonds to xylose residues of oligo- or polysaccharides. These bonds are known to exist in wood, and may exist in other woody plants and biofilms. The project employs the only enzyme known to break these bonds, XLE (xylan:lignin etherase) in order to create a peptide probe to indicate their presence. XLE’s homology to known xylosidases has permitted the identification of residues very likely to participate in the catalytic activity. These residues will be targets of modification that remove catalytic activity while retaining the ability to bind to XLE’s non-glycosidic substrate. The modifications will be performed using standard techniques of molecular biology and evaluated using with a novel indicating substrate for XLE activity. XLE’s homology with known glycosidases has also allowed identification of the enzyme’s potential binding site for its substrate. A peptide with an attached visible marker corresponding to this sequence but including the sequence changes that remove the catalytic activity will be synthesized. The resulting peptide will be tested to see if it reacts with wood pulp and biofilms. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.