Flow-microcalorimetric determination of enzymatic activities of the Trichoderma viridae-cellulase complex

Abstract

A method for measuring enzymatic activities of the Triehoderma viridae cellulase complex is described. The LKB 2277 Thermal activity monitor and a flow-mix mode were used.Enzymatic activities of samples of a crude cellulose complex have been determined using three substrates: cellobiose, carboxymethyl cellulose (CMC) and xylan. Some kinetic constants for cellobiase activity have been evaluated from the obtained results. A process of the endproduct inhibition of eellobiase resp. carboxymethyl eellulase activity by glucose has been observed too.The described method allows a direct determination of various enzymatic activities of the eellulase complex. Because of the high sensitivity and the simplicity, the method is a very suitable tool for studying the eeUulase complex, determining the optimal conditions of enzymatic break-down of the cellulosic materials and observing various mechanisms of the feedback control by products of enzymatic action. The procedure is completely general in nature and is applicable to other enzymatic systems. 9. /

◾ An Account in Calorimetric Research: From Fundamentals to Pharmaceuticals

“…This was noted in the aforementioned study by Beran and Paulicek (1992), who studied the activity of cellulases against macromolecular substrates. In this work, the calorimetric experiments only included soluble substrates such as carboxymethyl cellulose, but the broader potential of the method was discussed.…”

“…This application of calorimetry appears to hold a significant potential, but it has only been sporadically exploited. One early example includes the work by Beran and Paulicek (1992), who outlined how isothermal calorimetry could be used to assay enzyme kinetics on macromolecular substrates. More recently, Lonhienne et al (2001) made a more thorough calorimetric study of chitinases acting on insoluble chitin.…”

Kinetics of Enzymatic High-Solid Hydrolysis of Lignocellulosic Biomass Studied by Calorimetry

“…Evaluation of the Calorimetric Assay Calorimetry has been previously proven successful to characterize the activity of enzymes in dilute aqueous solutions [31][32][33], in complex crowded systems [34,35], and water-rich suspensions of insoluble subtrates [36,37] including hydrolysis of lignocellulose [19]. In all of these studies, calorimetry has been applied in the "titration mode", where enzyme and substrate are mixed by injections under continuous stirring.…”

Kinetic study of hydrogen peroxide decomposition by catalase in a flow-mix microcalorimetric system

“…Further thermodynamic research has been focused on characterising ligand-protein interactions [8], studying the formation of inclusion complexes [9], and analysing the transfer of solutes between two immiscible phases [10]. By contrast, only a few studies have been published on the use of flow-mix calorimetry for kinetic investigations [11][12][13]. This is rather surprising, since this tech-…”