288888 Neutron Reflectometry, QCM-D , and TIRF Study of the Interaction of Endoglucanases with Films of Amorphous Cellulose

Thursday, November 1, 2012: 8:55 AM
335 (Convention Center )
Michael Kent, Joint BioEnergy Institute, Sandia National Laboratory, Emeryville, CA

Cellulase enzyme cocktails include exoglucanases that digest cellulose chain ends and endoglucanases that cleave randomly at interior points along the chains.  While it is known that these enzymes work synergistically, the details are not fully understood.  In addition, cellulose binding domains (CBDs) are known to play an important role in the digestion of crystalline cellulose but much less is known about the benefit of CBDs in the digestion of amorphous cellulose.  Amorphous cellulose is of interest as pretreatment of biomass with ionic liquids, a promising next generation technology, results in a combination of amorphous cellulose and cellulose II.  Determining the full effects of endoglucanase activity is challenging because these enzymes can alter the structure of insoluble cellulose in addition to releasing soluble oligomers.  To unravel the actions of endoglucanases and the role of cellulose binding domains in enhancing activity on amorphous cellulose, we have combined studies of the profile of water through cellulose films during digestion by neutron reflectivity, measurements of changes in mass and film stiffness using a quartz crystal microbalance (QCM), and visualization of the motion of individual enzymes by total internal reflection fluorescence (TIRF) microscopy.  A trend has emerged in that the enzymes possessing a CBM penetrate into and digest within the bulk of the films to a much greater extent than those lacking a CBM.  In addition, much different results were observed for a classic endoglucanase and a processive endoglucanase that both possess CBMs, suggesting that penetration is driven by different mechanisms in the two cases.

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See more of this Session: Advances In Biofuels: DOE Bioenergy Research Centers I
See more of this Group/Topical: Sustainable Engineering Forum