title: Improved heterologous protein production by a tripeptidyl peptidase gene (AosedD) disruptant of the filamentous fungus Aspergillus oryzae.
abstract: Proteolytic degradation is one of the serious bottlenecks limiting the yields of heterologous protein production by Aspergillus oryzae. In this study, we selected a tripeptidyl peptidase gene AosedD (AO090166000084) as a candidate potentially degrading the heterologous protein, and performed localization analysis of the fusion protein AoSedD-EGFP in A. oryzae. As a result, the AoSedD-EGFP was observed in the septa and cell walls as well as in the culture medium, suggesting that AoSedD is a secretory enzyme. An AosedD disruptant was constructed to investigate an effect of AoSedD on the production level of heterologous proteins and protease activity. Both of the total protease and tripeptidyl peptidase activities in the culture medium of the AosedD disruptant were decreased as compared to those of the control strain. The maximum yields of recombinant bovine chymosin (CHY) and human lysozyme (HLY) produced by the AosedD disruptants showed approximately 2.9- and 1.7-fold increases, respectively, as compared to their control strains. These results suggest that AoSedD is one of the major proteases involved in the proteolytic degradation of recombinant proteins in A. oryzae.
authors.affiliation: Department of Biotechnology, The University of Tokyo, Japan.
authors.forename: Lin
authors.initials: L
authors.lastname: Zhu
authors.forename: Takeshi
authors.initials: T
authors.lastname: Nemoto
authors.forename: Jaewoo
authors.initials: J
authors.lastname: Yoon
authors.forename: Jun-ichi
authors.initials: J
authors.lastname: Maruyama
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Kitamoto
mesh_terms: D001236:Aspergillus oryzae / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D002473:Cell Wall / Q000201:enzymology; D003470:Culture Media / Q000737:chemistry; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000172:deficiency* / Q000378:metabolism; D055786:Gene Knockout Techniques; D060847:Metabolic Engineering; D011994:Recombinant Proteins / Q000096:biosynthesis*
chemical_list: D003470:Culture Media; D011994:Recombinant Proteins; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases