title: Quantifying Synergy, Thermostability, and Targeting of Cellulolytic Enzymes and Cellulosomes with Polymerization-Based Amplification.
abstract: We present a polymerization-based assay for determining the potency of cellulolytic enzyme formulations on pretreated biomass substrates. Our system relies on monitoring the autofluorescence of cellulose and measuring the attenuation of this fluorescent signal as a hydrogel consisting of poly(ethylene glycol) (PEG) polymerizes on top of the cellulose in response to glucose produced during saccharification. The one-pot method we present is label-free, rapid, highly sensitive, and requires only a single pipetting step. Using model enzyme formulations derived from Trichoderma reesei, Trichoderma longibrachiatum, Talaromyces emersonii and recombinant bacterial minicellulosomes from Clostridium thermocellum, we demonstrate the ability to differentiate enzyme performance based on differences in thermostability, cellulose-binding domain targeting, and endo/exoglucanase synergy. On the basis of its ease of use, we expect this cellulase assay platform to be applicable to enzyme screening for improved bioconversion of lignocellulosic biomass.
authors.affiliation: Lehrstuhl für Angewandte Physik and Center for Nanoscience, Ludwig-Maximilians-Universität, 80799 Munich, Germany.
authors.forename: Klara H
authors.initials: KH
authors.lastname: Malinowska
authors.affiliation: Lehrstuhl für Angewandte Physik and Center for Nanoscience, Ludwig-Maximilians-Universität, 80799 Munich, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Rind
authors.affiliation: Lehrstuhl für Angewandte Physik and Center for Nanoscience, Ludwig-Maximilians-Universität, 80799 Munich, Germany.
authors.forename: Tobias
authors.initials: T
authors.lastname: Verdorfer
authors.affiliation: Lehrstuhl für Angewandte Physik and Center for Nanoscience, Ludwig-Maximilians-Universität, 80799 Munich, Germany.
authors.forename: Hermann E
authors.initials: HE
authors.lastname: Gaub
authors.affiliation: Lehrstuhl für Angewandte Physik and Center for Nanoscience, Ludwig-Maximilians-Universität, 80799 Munich, Germany.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Nash
mesh_terms: D018533:Biomass; D002480:Cellulase / Q000378:metabolism*; D002482:Cellulose / Q000737:chemistry / Q000378:metabolism; D044603:Cellulosomes / Q000378:metabolism*; D048013:Acetivibrio thermocellus / Q000378:metabolism; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000737:chemistry / Q000378:metabolism; D011092:Polyethylene Glycols / Q000737:chemistry / Q000378:metabolism; D058105:Polymerization; D013059:Spectrum Analysis, Raman; D013696:Temperature; D014242:Trichoderma / Q000378:metabolism; D001617:beta-Glucosidase / Q000378:metabolism
chemical_list: D020136:Hydrogel, Polyethylene Glycol Dimethacrylate; D011092:Polyethylene Glycols; D002482:Cellulose; D001617:beta-Glucosidase; D002480:Cellulase