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Double disruption of the proteinase genes, tppA and pepE, increases the production level of human lysozyme by Aspergillus oryzae

  • Applied Genetics and Molecular Biotechnology
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Abstract

In this study, we investigated the effects of proteinase gene disruption on heterologous protein production by Aspergillus oryzae. The human lysozyme (HLY) was selected for recombinant production as a model for the heterologous protein. A tandem HLY construct fused with α-amylase (AmyB) was expressed by A. oryzae in which the Kex2 cleavage site was inserted at the upstream of HLY. HLY was successfully processed from AmyB and produced in the medium. We performed a systematic disruption analysis of five proteinase genes (pepA, pepE, alpA, tppA, and palB) in the HLY-producing strain with the adeA selectable marker. Comparative analysis indicated that disruption of the tppA gene encoding a tripeptidyl peptidase resulted in the highest increase (36%) in the HLY production. We further deleted the tppA gene in the pepE or palB disruptant with another selectable marker, argB. Consequently, a double disruption of the tppA and pepE genes led to a 63% increase in the HLY production compared to the control strain. This is the first study to report that the double disruption of the tppA and pepE genes improved the production level of a heterologous protein by filamentous fungi.

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Acknowledgment

This study was supported in part by the Program for Promotion of Basic Research Activities for Innovative Biosciences (PROBRAIN of Japan).

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Authors and Affiliations

  1. Department of Biotechnology, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo, 113-8657, Japan

    Feng Jie Jin, Taisuke Watanabe, Praveen Rao Juvvadi, Jun-ichi Maruyama, Manabu Arioka & Katsuhiko Kitamoto

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  1. Feng Jie Jin
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  2. Taisuke Watanabe
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  3. Praveen Rao Juvvadi
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  4. Jun-ichi Maruyama
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  5. Manabu Arioka
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  6. Katsuhiko Kitamoto
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Correspondence to Katsuhiko Kitamoto.

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Jin, F.J., Watanabe, T., Juvvadi, P.R. et al. Double disruption of the proteinase genes, tppA and pepE, increases the production level of human lysozyme by Aspergillus oryzae . Appl Microbiol Biotechnol 76, 1059–1068 (2007). https://doi.org/10.1007/s00253-007-1088-4

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