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Applied and Environmental Microbiology, June 2009, p. 4149-4154, Vol. 75, No. 12
0099-2240/09/$08.00+0     doi:10.1128/AEM.00318-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Regulation of the Display Ratio of Enzymes on the Saccharomyces cerevisiae Cell Surface by the Immunoglobulin G and Cellulosomal Enzyme Binding Domains{triangledown}

Junji Ito,1 Akihiko Kosugi,2 Tsutomu Tanaka,3 Kouichi Kuroda,4 Seiji Shibasaki,5 Chiaki Ogino,1 Mitsuyoshi Ueda,4 Hideki Fukuda,3 Roy H. Doi,6 and Akihiko Kondo1*

Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodaicho, Nada-ku, Kobe 657-8501, Japan,1 Japan International Research Center for Agricultural Sciences (JIRCAS), Tsukuba, Ibaraki 305-8686, Japan,2 Organization of Advanced Science and Technology, Kobe University, 1-1 Rokkodaicho, Nada-ku, Kobe 657-8501, Japan,3 Department of Applied Biochemistry, Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwake-cho, Sakyo-ku, Kyoto 606-8502, Japan,4 Department of Pharmacy, School of Pharmacy, Hyogo University of Health Sciences, Minatojima, Chuo-ku, Kobe, 650-8530, Japan,5 Section of Molecular and Cellular Biology, University of California, Davis, California 95616-85356

Received 9 February 2009/ Accepted 20 April 2009

We constructed a novel cell surface display system to control the ratio of target proteins on the Saccharomyces cerevisiae cell surface, using two pairs of protein-protein interactions. One protein pair is the Z domain of protein A derived from Staphylococcus aureus and the Fc domain of human immunoglobulin G. The other is the cohesin (Coh) and dockerin (Dock) from the cellulosome of Clostridium cellulovorans. In this proposed displaying system, the scaffolding proteins (fusion proteins of Z and Coh) were displayed on the cell surface by fusing with the 3' half of {alpha}-agglutinin, and the target proteins fused with Fc or Dock were secreted. As a target protein, a recombinant Trichoderma reesei endoglucanase II (EGII) was secreted into the medium and immediately displayed on the yeast cell surface via the Z and Fc domains. Display of EGII on the cell surface was confirmed by hydrolysis of β-glucan as a substrate, and EGII activity was detected in the cell pellet fraction. Finally, two enzymes, EGII and Aspergillus aculeatus β-glucosidase 1, were codisplayed on the cell surface via Z-Fc and Dock-Coh interactions, respectively. As a result, the yeast displaying two enzymes hydrolyzed β-glucan to glucose very well. These results strongly indicated that the proposed strategy, the simultaneous display of two enzymes on the yeast cell surface, was accomplished by quantitatively controlling the display system using affinity binding.

* Corresponding author. Mailing address: Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodaicho, Nada-ku, Kobe 657-8501, Japan. Phone and fax: 81-78-803-6196. E-mail: akondo{at}kobe-u.ac.jp

{triangledown} Published ahead of print on 1 May 2009.

Applied and Environmental Microbiology, June 2009, p. 4149-4154, Vol. 75, No. 12
0099-2240/09/$08.00+0     doi:10.1128/AEM.00318-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Tsai, S.-L., Oh, J., Singh, S., Chen, R., Chen, W. (2009). Functional Assembly of Minicellulosomes on the Saccharomyces cerevisiae Cell Surface for Cellulose Hydrolysis and Ethanol Production. Appl. Environ. Microbiol. 75: 6087-6093 [Abstract] [Full Text]  


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