Document Detail


Display of active enzymes on the cell surface of Escherichia coli using PgsA anchor protein and their application to bioconversion.
MedLine Citation:
PMID:  16133338     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
We have developed a novel Escherichia coli cell surface display system by employing PgsA as an anchoring motif. In our display system, C-terminal fusion to PgsA anchor protein from Bacillus subtilis was used. The enzymes selected for display were alpha-amylase (AmyA) from Streptococcus bovis 148 and lipase B (CALB) from Candida antarctica. The molecular mass values of AmyA and CALB are approximately 77 and 34 kDa, respectively. The enzymes were displayed on the surface as a fusion protein with a FLAG peptide tag at the C terminus. Both the PgsA-AmyA-FLAG and PgsA-CALB-FLAG fusion proteins were shown to be displayed by immunofluorescence labeling using anti-FLAG antibody. The displayed enzymes were active forms, and AmyA and CALB activities reached 990 U/g (dry cell weight) and 4.6 U/g (dry cell weight), respectively. AmyA-displaying E. coli cells grew utilizing cornstarch as the sole carbon source, while CALB-displaying E. coli cells catalyzed enantioselective transesterification, indicating that they are effective whole-cell biocatalysts. Since a target enzyme with a size of 77 kDa and an industrially useful lipase have been successfully displayed on the cell surface of E. coli for the first time, PgsA protein is probably a useful anchoring motif to display various enzymes.
Authors:
Junya Narita; Kenji Okano; Toshihiro Tateno; Takanori Tanino; Tomomitsu Sewaki; Moon-Hee Sung; Hideki Fukuda; Akihiko Kondo
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2005-08-20
Journal Detail:
Title:  Applied microbiology and biotechnology     Volume:  70     ISSN:  0175-7598     ISO Abbreviation:  Appl. Microbiol. Biotechnol.     Publication Date:  2006 May 
Date Detail:
Created Date:  2006-04-07     Completed Date:  2006-08-21     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  8406612     Medline TA:  Appl Microbiol Biotechnol     Country:  Germany    
Other Details:
Languages:  eng     Pagination:  564-72     Citation Subset:  IM    
Affiliation:
Division of Molecular Science, Graduate School of Science and Technology, Kobe University, Nada-ku, Kobe, 657-8501, Japan.
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MeSH Terms
Descriptor/Qualifier:
Candida / enzymology
Catalysis
Cell Membrane / metabolism*
Enzymes, Immobilized / genetics,  metabolism*
Escherichia coli / cytology*,  metabolism*
Gene Expression
Lipase / genetics,  metabolism
Membrane Proteins / genetics,  metabolism*
Recombinant Proteins
Transferases (Other Substituted Phosphate Groups) / genetics,  metabolism*
alpha-Amylases / genetics,  metabolism
Chemical
Reg. No./Substance:
0/Enzymes, Immobilized; 0/Membrane Proteins; 0/Recombinant Proteins; EC 2.7.8.-/Transferases (Other Substituted Phosphate Groups); EC 2.7.8.5/CDP-diacylglycerol-glycerol-3-phosphate 3-phosphatidyltransferase; EC 3.1.1.-/lipase B, Candida antarctica; EC 3.1.1.3/Lipase; EC 3.2.1.1/alpha-Amylases

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