The CellG5 cellulase reagent used in the K-CellG5 Assay Kit represents a significant improvement on the CellG3 reagent (K-CellG3 Assay Kit) that was introduced by Megazyme in 2012. The CellG5 reagent employs high purity thermostable β-glucosidase and blocked 4-nitrophenyl-β-D-cellopentaoside (BPNPG5) containing a cellopentaose core and exhibits vastly improved sensitivity for some cellulases. In addition, the exchange of the benzylidene blocking group in CellG3 for 3-keto-butylidene in CellG5 improves the substrate’s water solubility significantly, allowing for a reduction in the concentration of DMSO required in the assay. As DMSO is known to inhibit certain cellulases, this is another benefit in using CellG5.
Megazyme recommends the use the K-CellG5 Assay Kit for all assays for the measurement of endo-cellulase.
K-CellG5 Assay Principle:
On hydrolysis of BPNPG5 by cellulase, the 4-nitrophenyl-β-D-cellooligosaccharide fragment is immediately hydrolysed to D-glucose and free 4-nitrophenol (pNP) by the β-glucosidase present in the reagent mixture. Thus, the rate of release of pNP relates directly to the rate of hydrolysis of BPNPG5 by cellulase. The reaction is stopped and the phenolate colour is developed by addition of Tris buffer solution (pH 10).