Expression of Active Calf Chymosin in Aspergillus Niger

摘要

Calf chymosin is an important protease used widely in the industry of food, medicine and plastics. We try use Aspergillus niger as a genetically engineered host to produce the heterologous protein. The prochymosin cDNA was cloned from the abomasum dells of unweaned calves by reverse transcription polymerase chain reaction (RT-PCR). The cloned fragment of 1100 bp in vector pUCm-T was then subcloned into an expression vector pyG1.2 containing the pyrG gene. The insert fragment was fused in frame with glucoamylase coding region under the control of A. niger glaA promoter. The recombined plasmid containing prochymosin cDNA transformed the M54, a pyrG gene auxotroph of Aspergillus niger. Three transformants was selected and they showed no obvious difference in expression and secretion of chymosin. The result of SDS-PAGE indicated that there was protein band of 36KD with the same molecular weight as chymosin. The milk coagulation test proved that the active chymosin had been expressed and secreted successfully by the recombinant A.niger. This study further emphasizes the potential importance of aspergillus niger as a useful host for the production of heterologous proteins with authentic biological properties.