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Thermal denaturation ofTrichoderma reesei cellulases studied by differential scanning calorimetry and tryptophan fluorescence

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Abstract

The thermal denaturation of four purified Trichoderma reesei cellulase components, cellobiohydrolase (CBH) I, CBH II, endoglucanase (EG) I, and EG II, has been monitored using a combination of classical temperature/activity profiles, differential scanning calorimetry (DSC), and thermal scanning fluorescence emission spectrometry. Significant correlations were found between the results of enzyme activity studies and the results obtained through the more direct physical approaches, in that both DSC and the activity studies showed EG II (Tm = 75°C) to be much more thermostable (by 10–11 °C) than the other three enzymes, all three of which were shown by both activity profiles and DSC to be very similar in thermal stability. The temperature dependence of the wavelength of maximum tryptophan emission showed a parallel result, with the three enzymes exhibiting less thermostable activity being grouped together in this regard, and EG II differing from the other three in maintaining a less-exposed tryptophan microenvironment at temperatures as high as 73 °C. The DSC results suggested that at least two transitions are involved in the unfolding of each of the cellulase components, the first (lower-temperature) of which may be the one correlated with activity loss.

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Authors and Affiliations

  1. Biotechnology Research Branch, Fuels and Chemicals Research and Engineering Division, Solar Energy Research Institute, 80401, Golden, CO

    J. O. Baker, K. Tatsumoto, K. Grohmann & M. E. Himmel

  2. Chemical Technology Division, Oak Ridge National Laboratory, 37811-6194, OakRidge, TN

    J. Woodward & J. M. Wichert

  3. Department of Food Science & Technology, University of California, 95616, Davis, CA

    S. P. Shoemaker

Authors
  1. J. O. Baker
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  2. K. Tatsumoto
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  3. K. Grohmann
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  4. J. Woodward
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  5. J. M. Wichert
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  6. S. P. Shoemaker
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  7. M. E. Himmel
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Baker, J.O., Tatsumoto, K., Grohmann, K. et al. Thermal denaturation ofTrichoderma reesei cellulases studied by differential scanning calorimetry and tryptophan fluorescence. Appl Biochem Biotechnol 34, 217–231 (1992). https://doi.org/10.1007/BF02920547

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  • DOI: https://doi.org/10.1007/BF02920547

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