Abstract
The interaction between cellulase enzymes and their substrates is of central importance to several technological and scientific challenges. Here we report that the binding of cellulose binding modules (CBM) from Trichoderma reesei cellulases Cel6A and Cel7A show a major difference in how they interact with substrates originating from wood compared to bacterial cellulose. We found that the CBM from TrCel7A recognizes the two substrates differently and as a consequence shows an unexpected way of binding. We show that the substrate has a large impact on the exchange rate of the studied CBM, and moreover, CBM-TrCel7A seems to have an additional mode of binding on wood derived cellulose but not on cellulose originating from bacterial source. This mode is not seen in double CBM (DCBM) constructs comprising both CBM-TrCel7A and CBM-TrCel6A. The linker length of DCBMs affects the binding properties, and slows down the exchange rates of the proteins and thus, can be used to analyze the differences between the single CBM. These results have impact on the cellulase research and offer new understanding on how these industrially relevant enzymes act.
Highlights
The interaction between cellulase enzymes and their substrates is of central importance to several technological and scientific challenges
Double Carbohydrate binding modules (CBMs) (DCBMs) with different linker lengths were produced as HFBI hydrophobin fusions (HFBI-double CBM (DCBM)) to aid purification (DCBM sequences shown in Supplementary Figure S1) as described previously[19]
It is especially notable that the synergies between modules seen for the DCBM-12 and DCBM-24 when binding to Bacterial microcrystalline cellulose (BMCC) are not seen when the proteins bind to Cellulose nanofbrils (CNF)
Summary
The interaction between cellulase enzymes and their substrates is of central importance to several technological and scientific challenges. The linker length of DCBMs affects the binding properties, and slows down the exchange rates of the proteins and can be used to analyze the differences between the single CBM These results have impact on the cellulase research and offer new understanding on how these industrially relevant enzymes act. In this work we focus on the type of CBMs that are found in fungi and are classified as family[1,2,8,9] These have been the subject of much investigation because of the efficiency and importance of fungal enzyme systems for degrading cellulosic material both as a part of the ecosystem and increasingly for technical applications[10,11]. Using a set of differently designed molecules, different sources of cellulose, and accurate measurement techniques enabling the study of binding kinetics allowed a new set of tools to investigate cellulose as complex structure and the structure-function relationship of the enzymes acting on it
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