A Low-Volume, Parallel Copper-Bicinchoninic Acid (BCA) Assay for Glycoside Hydrolases.

The quantitation of liberated reducing sugars by the copper-bicinchoninic acid (BCA) assay provides a highly sensitive method for the measurement of glycoside hydrolase (GH) activity, particularly on soluble polysaccharide substrates. Here we describe a straightforward method adapted to low-volume polymerase chain reaction (PCR) tubes that enables the rapid, parallel determination of GH kinetics in applications ranging from initial activity screening and assay optimization to precise Michaelis-Menten analysis.

Configured for the Human Gut Microbiota: Molecular Mechanisms of Dietary ß-Glucan Utilization.

The ß-glucans are a disparate group of structurally diverse polysaccharides, whose members are widespread in human diets as components of the cell walls of plants, algae, and fungi (including yeasts), and as bacterial exopolysaccharides. Individual ß-glucans from these sources have long been associated with positive effects on human health through metabolic and immunological effects. Remarkably, the ß-configured glucosidic linkages that define these polysaccharides render them inaccessible to the limited repertoire of digestive enzymes encoded by the human genome. As a result, the various ß-glucans become fodder for the human gut microbiota (HGM) in the lower gastrointestinal tract, where they influence community composition and metabolic output, including fermentation to short chain fatty acids (SCFAs). Only recently, however, have the specific molecular systems that enable the utilization of ß-glucans by select members of the HGM been fully elucidated by combined genetic, biochemical, and structural biological approaches. In the context of ß-glucan structures and their effects on human nutrition and health, we summarize here the functional characterization of individual polysaccharide utilization loci (PULs) responsible for the saccharification of mixed-linkage ß(1→3)/ß(1→4)-glucans, ß(1→6)-glucans, ß(1→3)-glucans, ß(1→2)-glucans, and xyloglucans in symbiotic human gut bacteria. These exemplar PULs serve as well-defined biomarkers for the prediction of ß-glucan metabolic capability in individual bacterial taxa and across the global human population.