Domain replacement to elucidate the role of B domain in CGTase thermostability and activity

► We attempted to improve CGTase thermostability by whole B domain engineering. ► Whole B domain was substituted and the approach has never reported before. ► A mutant exhibited 2.5-fold thermostability enhancement. ► Improvement may be cause by better structural packing and molecular interactions. ► Five mutants lost activity conceivably due to substrate-binding cleft distortion.