TY - JOUR
T1 - Thermal Denaturation of Trichoderma Reesei Cellulases Studied by Differential Scanning Calorimetry and Tryptophan Fluorescence Shifts
AU - Baker, J. O.
AU - Tatsumoto, K.
AU - Grohmann, K.
AU - Woodward, J.
AU - Wichert, J. M.
AU - Shoemaker, S. P.
AU - Himmel, M. E.
PY - 1992
Y1 - 1992
N2 - 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.
AB - 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.
KW - T. reesei cellulases
KW - Thermal protein denaturation
UR - http://www.scopus.com/inward/record.url?scp=0000288652&partnerID=8YFLogxK
U2 - 10.1007/BF02920547
DO - 10.1007/BF02920547
M3 - Article
AN - SCOPUS:0000288652
SN - 0273-2289
VL - 34-35
SP - 217
EP - 231
JO - Applied Biochemistry and Biotechnology - Part A Enzyme Engineering and Biotechnology
JF - Applied Biochemistry and Biotechnology - Part A Enzyme Engineering and Biotechnology
IS - 1
ER -