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Domain coupling in a multimodular cellobiohydrolase CbhA from Clostridium thermocellum
Author(s) -
Kataeva Irina A.,
Brewer John M.,
Uversky Vladimir N.,
Ljungdahl Lars G.
Publication year - 2005
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2005.06.074
Subject(s) - clostridium thermocellum , ethylenediaminetetraacetic acid , chemistry , cooperativity , thermophile , differential scanning calorimetry , biochemistry , crystallography , chromatography , enzyme , cellulase , organic chemistry , thermodynamics , physics , chelation
Cellobiohydrolase A (CbhA) from Clostridium thermocellum is composed of an N‐terminal carbohydrate‐binding domain 4 (CBD4), an immunoglobulin‐like domain (Ig), a glycoside hydrolase 9 (GH9), X1 1 and X1 2 domains, a CBD3, and a dockerin domain. All domains, except the Ig, bind Ca 2+ . The following constructs were made: X1 2 , X1 1 X1 2 , CBD3, X1 1 X1 2 ‐CBD3, Ig, GH9, Ig‐GH9, Ig‐GH9‐X1 1 X1 2 , and Ig‐GH9‐X1 1 X1 2 ‐CBD3. Interactions between domains in (1) buffer, (2) with Ca 2+ , or (3) ethylenediaminetetraacetic acid (EDTA) were studied by differential scanning calorimetry. Thermal unfoldings of all constructs were irreversible. Calcium increased T d and cooperativity of unfolding. Multi‐domain constructs exhibited more cooperative unfolding in buffer and in the presence of EDTA than did individual domains. They denatured by mechanism simpler than expected from their modular architecture. The results indicate that domain coupling in thermophilic proteins constitutes a significant stabilizing factor.