In vitro expression of Penicillium janthinellum cellobiohydrolase I gene in a coupled transcription-translation system

摘要

All enzymatically-active fungal cellobiohydrolase I (CBH I) was first synthesized in a coupled reticulocyte lysate system lacking of glycosylation moditication by the template DNA_(Cbhl) in the presence of T7 RNA polymerase. The synthesized CBH I had the expected size (57 kDa) and catalyzed the substrate of p-nitrophenyl-#beta#-D-cellobioside (pNPC), and had no activity against carboxymethyl cellulose (CMC-Na). The K_m and V_(max) values of the CBH I for pNPC were 0.82 mmol and 0.067 #mu#mol min~(-1) per lag enzyme, respectively. The results indicated that glycosylation may not be necessary for enzymatic activity of fungal cellulases.