Quantitative assessment of the enzymatic degradation of amorphous cellulose by using a quartz crystal microbalance with dissipation monitoring

M. Suchy, Markus Linder, Tekla Tammelin, J.M. Campbell, T. Vuorinen, E. Kontturi (Corresponding Author)

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Abstract

The systematic evaluation of the degradation of an amorphous cellulose film by a monocomponent endoglucanase (EG I) by using a quartz crystal microbalance with dissipation monitoring (QCM-D) identified several important aspects relevant to the study the kinetics of cellulose degradation by enzymes. It was demonstrated that, to properly evaluate the mechanism of action, steady state conditions in the experimental set up need to be reached. Rinsing or diluting the enzyme, as well as concentration of the enzyme, can have a pronounced effect on the hydrolysis. Quantification of the actual hydrolysis was carried out by measuring the film thickness reduction by atomic force microscopy after the enzymatic treatment. The values correlated well with the frequency data obtained by QCM-D measurement for corresponding films. This demonstrated that the evaluation of hydrolysis by QCM-D can be done quantitatively. Tuning of the initial thickness of films enabled variation of the volume of substrate available for hydrolysis which was then utilized in establishing a correlation between substrate volume and hydrolytic activity of EG I as measured by QCM-D. It was shown that, although the amount of substrate affects the absolute rate of hydrolysis, the relative rate of hydrolysis does not depend on the initial amount of substrate in steady state system. With this experimental setup it was also possible to demonstrate the impact of concentration on crowding of enzyme and subsequent hydrolysis efficiency. This effort also shows the action of EG I on a fully amorphous substrate as observed by QCM-D. The enzyme was shown to work uniformly within the whole volume of swollen film, however being unable to fully degrade the amorphous film.
Original languageEnglish
Pages (from-to)8819-8828
Number of pages10
JournalLangmuir
Volume27
Issue number14
DOIs
Publication statusPublished - 2011
MoE publication typeA1 Journal article-refereed

Fingerprint

Quartz crystal microbalances
quartz crystals
cellulose
Cellulose
microbalances
hydrolysis
Hydrolysis
dissipation
degradation
Degradation
enzymes
Monitoring
Enzymes
Substrates
Amorphous films
4 alpha-glucanotransferase
Cellulose films
Enzyme kinetics
crowding
Cellulase

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Suchy, M. ; Linder, Markus ; Tammelin, Tekla ; Campbell, J.M. ; Vuorinen, T. ; Kontturi, E. / Quantitative assessment of the enzymatic degradation of amorphous cellulose by using a quartz crystal microbalance with dissipation monitoring. In: Langmuir. 2011 ; Vol. 27, No. 14. pp. 8819-8828.
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abstract = "The systematic evaluation of the degradation of an amorphous cellulose film by a monocomponent endoglucanase (EG I) by using a quartz crystal microbalance with dissipation monitoring (QCM-D) identified several important aspects relevant to the study the kinetics of cellulose degradation by enzymes. It was demonstrated that, to properly evaluate the mechanism of action, steady state conditions in the experimental set up need to be reached. Rinsing or diluting the enzyme, as well as concentration of the enzyme, can have a pronounced effect on the hydrolysis. Quantification of the actual hydrolysis was carried out by measuring the film thickness reduction by atomic force microscopy after the enzymatic treatment. The values correlated well with the frequency data obtained by QCM-D measurement for corresponding films. This demonstrated that the evaluation of hydrolysis by QCM-D can be done quantitatively. Tuning of the initial thickness of films enabled variation of the volume of substrate available for hydrolysis which was then utilized in establishing a correlation between substrate volume and hydrolytic activity of EG I as measured by QCM-D. It was shown that, although the amount of substrate affects the absolute rate of hydrolysis, the relative rate of hydrolysis does not depend on the initial amount of substrate in steady state system. With this experimental setup it was also possible to demonstrate the impact of concentration on crowding of enzyme and subsequent hydrolysis efficiency. This effort also shows the action of EG I on a fully amorphous substrate as observed by QCM-D. The enzyme was shown to work uniformly within the whole volume of swollen film, however being unable to fully degrade the amorphous film.",
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Quantitative assessment of the enzymatic degradation of amorphous cellulose by using a quartz crystal microbalance with dissipation monitoring. / Suchy, M.; Linder, Markus; Tammelin, Tekla; Campbell, J.M.; Vuorinen, T.; Kontturi, E. (Corresponding Author).

In: Langmuir, Vol. 27, No. 14, 2011, p. 8819-8828.

Research output: Contribution to journalArticleScientificpeer-review

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AB - The systematic evaluation of the degradation of an amorphous cellulose film by a monocomponent endoglucanase (EG I) by using a quartz crystal microbalance with dissipation monitoring (QCM-D) identified several important aspects relevant to the study the kinetics of cellulose degradation by enzymes. It was demonstrated that, to properly evaluate the mechanism of action, steady state conditions in the experimental set up need to be reached. Rinsing or diluting the enzyme, as well as concentration of the enzyme, can have a pronounced effect on the hydrolysis. Quantification of the actual hydrolysis was carried out by measuring the film thickness reduction by atomic force microscopy after the enzymatic treatment. The values correlated well with the frequency data obtained by QCM-D measurement for corresponding films. This demonstrated that the evaluation of hydrolysis by QCM-D can be done quantitatively. Tuning of the initial thickness of films enabled variation of the volume of substrate available for hydrolysis which was then utilized in establishing a correlation between substrate volume and hydrolytic activity of EG I as measured by QCM-D. It was shown that, although the amount of substrate affects the absolute rate of hydrolysis, the relative rate of hydrolysis does not depend on the initial amount of substrate in steady state system. With this experimental setup it was also possible to demonstrate the impact of concentration on crowding of enzyme and subsequent hydrolysis efficiency. This effort also shows the action of EG I on a fully amorphous substrate as observed by QCM-D. The enzyme was shown to work uniformly within the whole volume of swollen film, however being unable to fully degrade the amorphous film.

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