Construction of a low-serine-type-carboxypeptidase-producing mutant of Aspergillus oryzae by the expression of antisense RNA and its use as a host for heterologous protein secretion.
Using an antisense control strategy, we isolated an Aspergillus oryzae mutant that produced low levels of carboxypeptidases (CPases). The mutant TFC-1 expressed the antisense RNA of the structural gene of CPase O and showed about 30% of the CPase activity in the parent strain. Gel filtration analysis indicated that this mutant decreased the CPase activities not only of CPase O but also of CPase O-1 and O-2. This result indicated that the antisense RNA was able to control the expression of the CPase genes as a group. Using the mutant as a heterologous protein expression host that produced the low levels of CPases, a stable and higher level of lysozyme expression could be obtained compared with the wild-type. In vitro proteolytic degradation assay also demonstrated that human lysozyme was degraded by purified CPase O.
Zheng
X F
XF
Faculty of Agriculture, Tokyo University of Agriculture and Technology, Japan.
Kobayashi
Y
Y
Takeuchi
M
M
D001236:Aspergillus oryzae / Q000235:genetics*; D015139:Blotting, Southern; D002268:Carboxypeptidases / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D009113:Muramidase / Q000096:biosynthesis*; D009154:Mutation; D016372:RNA, Antisense / Q000502:physiology*; D012694:Serine; D014170:Transformation, Genetic
D016372:RNA, Antisense; D012694:Serine; D009113:Muramidase; D002268:Carboxypeptidases