Industrial Uses of Thermophilic Cellulase
Protein Structure and Function (CHEM 645)
Spring 2004
University of Delaware
   


Introduction

   
    Thermophiles come from what was thought to be unihabitable places of the earth, such as steam vents, volcanic areas and arctic  places.  These small bacteria like organisms can withstand extreme temperatures.  The images below show a few places where thermophiles can be found.  The image on the left shows a hot steam vent under the ocean and the picture to the right shows a picture of vents at Sylvan Springs.

         

Because these thermophiles live in extremely hot temperatures, their enzymes can still be active at these high temperatures with out denaturing.  There have been numerous enzymes that have been extracted from thermophiles that are now being put to use in industry1-4.  Cellulose is one of the most abundant organic molecules on the earth and that is why cellulases are so important1-3.  Thermophilic enzymes are ideal for using because they can withstand high temperatures without being denatured like most other proteins would be1-3.   For industrial purposes, this means that the enzymes can still be active and keep the product sterilized because its being prepared at higher temperatures.  Also, its better for the environment to use thermophilic cellulases compared to chemicals1-3.The following is a list of processes that are used with thermophilic cellulase:

Food Processing

Textile Processes

Paper Processing

Ethanol Fuel Production



Food Processing
   
    One of the primary uses of thermophilic cellulases that I found in the food industry was for the use of preparing juices.  The example that I will use will be based on apple juice.  The juice used to be prepared by crushing the apples and then filtering the juice1.  However, this wasn't good for yield.  Now, the cellulases are added, after crushing, to the leftover pulp1.  By adding the cellulases, they are increasing the yield of the juice and preventing further contamination because they can perform the juicing process at higher temperatures1.




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References

Textile Processes

   
The textile industry uses thermophilic cellulases for of reasons including uses in detergeants for keeping color brightness for longer and creating the stone washed look in jeans1-3,11.  It can be seen that after numerous washings, clothes tend to have a faded and fuzzy look to them.  Detergent companies have started adding thermophilic cellulases to their products to decrease the discoloration and fuzzing effects caused by numerous washes2-3,11.  The following picture shows what happens to fibers after numerous washings with and without the use of the cellulases:



As you can see from the pictures above, there is a lot more degreadation occuring to the fibers after washing without the use of the cellulases.
   
    One of the other textile processes that commonly uses thermophilic cellulases is the stone washing of jeans to make them appear faded and afterwards the jeans tend to be softer.  At first, to get the stoned washed look, the new jeans would be washed with pumice stones that would break off the dye from the surface of the jeans and cause the threads to degrade1.   After washing with the stones, they would have to manually be removed from the jeans, as well as, the stones creating a powder that would be harmful to the people that had to remove them1.   Now they add thermophilic cellulases in the wash with the jeans and the cellulases only eataway at the dye that is used to color the jeans and it maintains the durability1-3,11.  They create different affects depending on the amount of the cellulases they added to the wash.  Besides limiting the pollution that is created, the jeans can be stoned washed in less time than washing with the stones2.
 

    You might be thinking how can the cellulases brighten and keep color and then they are able to make it fade?  Well, if you looked at the general overview of cellulases, I mentioned the three different types.   Well, if you change the ratio of these types of cellulases, you can get different types of reactions.

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Paper Processing

    To create and recycle paper strong chemicals are used that are both unsafe to us and the environment9-10.   Because paper is mad out of wood, it obvious that it's more difficult to break it down in to small pieces that can made into paper because of the cellulose cell walls in the wood.  Well cellulases would be good for doing this without using harsh chemicals that can harm the environment1.  One of the big processes that they are used for is the bleaching process.  They use to use chlorine based bleaching chemicals that would be harmful not only to the environment but people's health as well2,9-10.  Now, by adding the thermophilic cellulases, they bleach the wood pulp to make it white.  And it is also used to take the ink out of recycled paper.  Both of these ways do not degrade the strength of the fiber.  The picture below shows the effects of using chemicals that have been used and the fibers after cellulase application:




Notice that the chemically treated fiber (left picture) is much thinner and has more frayed ends coming off of it than the fibers that were treated with the thermophilic cellulases10.   As you can see, thermophilic cellulases tend to be a better alternative to what is done with chemicals when doing paper processing.  Not only do they keep fiber strength but they are more environmentally friendly than the chemicals that were used. 

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References


Ethanol Fuel Production
   
    One of the most recent developments for the use of thermophilic cellulases has been the production of ethanol for fuel.  Iogen Corporation is the forerunner for this type of production with a process they call EcoEthanol.  The great thing about this process is that uses the non-edible portions of corn and wheat8.  Currently ethanol fuel is being made from corn, however, this limits the animal feed that is produced.  By using EcoEthanol, production of CO2 will be decreased by 90% with comparison to gasoline, which is a lot better than ethanol fuel that is produced from corn8.  Iogen is currently working on a plant for mass production.

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References


A Brief Overview of Cellulase
   
   
Cellulase is the enzyme that is used to break up cellulose into glucose or other oligosacharide compounds.  Cellulose is composed of glucose units linked together by a ß-1.4 glycosidic bond.  The picture below shows the basic structure of a cellulose molecule.  The actual number of units of glucoses can be numerous and is not defined to a specific number (it can be up in the thousands).



Cellulases are defined by their ability to cut the
ß-1.4 glycosidic bond.  There are three different types of cellulases, which are explained below5:



By varying how much of each type goes into a mixture, different types of reactions can occur.   Below is a picture of an endoglucanase structure from Rhodothermus marinus Cel12A with a cellulose in the binding domain of the Cellulose6:


Cellulases tend to have three main components to them.  The cellulose binding domain or CBD is responsible for bringing the molecule into position with respect to the the cellulose molecule7.  The catalytic core is where the cellulose is broken down and the flexible and heavily glycosylated linker region guides the CBD and the core over the substrate7.

For more information on Cellulases, I such you visit this webpage:  Microbial Cellulose Utilization: Fundamentals and Biotechnology

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References

1.  M.K. Bhat.  Cellulases and related enzymes in biotechnology.  Biotechnology Advances.  2000, 18, 355-383.

2.  Haki, G.D.; Rakshit, S.K.  Developments in industrially important thermostable enzymes: a review.  Bioresource Review.  2003, 89, 17-34.

3.  Vielle, C.; Zeikus, G.J.  Hyperthermophilic Enzymes: Sources, Uses, and Molecular Mechanisms for Thermostability.  Microbiology and Molecular Biology Reviews.  2001, 65(1), 1-34.

4.  Schiraldi, C.; De Rosa,M.  The production of biocatalysts and biomolecules from extremophiles.  Trends in Biotechnology.  2002, 20(12), 515-521.

5.  Hreggvidsson, G.O.; Kaiste, E.; Holst, O.; Eggertsson, G.; Palsdottier, A.; Kristjansson, J.K.  An Extremely Thermostable Cellulase from the Thermophilic Eubacterium Rhodothermus marinus.  Applied and Environmental Microbiology.  1996, 62(8), 3047-3049.

6.  Crennell, S.J.; Hreggvidsson, G.O.; Karisson, E.N.  The Structure of Rhodothermus marinus Cel12A, A Highly Thermostable Family 12 Endoglucanase, at 1.8Å Resolution. J. Mol. Biol.  2002, 320, 883-897.

7.  Hirvonen, M.; Papageorgiou, A.C.  Crystal Structure of a Family 45 Endoglucanases from Melanocarpus albomyces:  Mechanist Implications Based on the Free and Cellobiose-bound Forms.  J. Mol. Biol.  2003, 329, 403-410.

8.  Iogen doubles EcoEthanol Capacity.  April 28, 2003. http://www.iogen.ca/news/28_03_2003.html (accessed May 17, 2003).

9.  Pelach, M.A.; Pastor, F.J.; Puig, J.; Vilaseca, F.; Mutje, P.  Enzymic deinking
of old newspapers with cellulase. Process Biochemistry. 2003, 38, 1063-1067.

10.  Dienes, D.; Egyhazi, A.; Reczey, K.  Treatment of recycled fiber with
Trichoderma cellulases.  Industrial Crops and Products.  2004, article in press.

11.  Csiszar, E.; Losonczi, A. Szakacs, G. Rusznak, I.; Bezur, L.; Reicher, J.  Enzymes and chelating agent in cotton pretreatment.  Journal of Biotechnology.  2001, 89, 271-279.


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