Cell-Surface display of heterologous proteins: From high-throughput screening to environmental applications

2002 ◽  
Vol 79 (5) ◽  
pp. 496-503 ◽  
Author(s):  
Wilfred Chen ◽  
George Georgiou
Keyword(s):  
Cell Surface ◽  
High Throughput ◽  
High Throughput Screening ◽  
Surface Display ◽  
Cell Surface Display ◽  
Environmental Applications ◽  
Heterologous Proteins

Construction of a novel system for cell surface display of heterologous proteins on Pichia pastoris

2007 ◽  
Vol 29 (10) ◽  
pp. 1561-1566 ◽  
Author(s):  
Qingjie Wang ◽  
Lei Li ◽  
Min Chen ◽  
Qingsheng Qi ◽  
Peng George Wang
Keyword(s):  
Pichia Pastoris ◽  
Cell Surface ◽  
Surface Display ◽  
Cell Surface Display ◽  
Heterologous Proteins

scholarly journals Functional Cell Surface Display and Controlled Secretion of Diverse Agarolytic Enzymes by Escherichia coli with a Novel Ligation-Independent Cloning Vector Based on the Autotransporter YfaL

2012 ◽  
Vol 78 (9) ◽  
pp. 3051-3058 ◽  
Author(s):  
Hyeok-Jin Ko ◽  
Eunhye Park ◽  
Joseph Song ◽  
Taek Ho Yang ◽  
Hee Jong Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Fluorescent Protein ◽  
Surface Display ◽  
Cell Surface Display ◽  
Heterologous Proteins ◽  
Display System ◽  
Reporter Protein ◽  
Content Type ◽  
Ligation Independent Cloning

ABSTRACTAutotransporters have been employed as the anchoring scaffold for cell surface display by replacing their passenger domains with heterologous proteins to be displayed. We adopted an autotransporter (YfaL) ofEscherichia colifor the cell surface display system. The critical regions in YfaL for surface display were identified for the construction of a ligation-independent cloning (LIC)-based display system. The designed system showed no detrimental effect on either the growth of the host cell or overexpressing heterologous proteins on the cell surface. We functionally displayed monomeric red fluorescent protein (mRFP1) as a reporter protein and diverse agarolytic enzymes fromSaccharophagus degradans2-40, including Aga86C and Aga86E, which previously had failed to be functional expressed. The system could display different sizes of proteins ranging from 25.3 to 143 kDa. We also attempted controlled release of the displayed proteins by incorporating a tobacco etch virus protease cleavage site into the C termini of the displayed proteins. The maximum level of the displayed protein was 6.1 × 104molecules per a single cell, which corresponds to 5.6% of the entire cell surface of actively growingE. coli.

scholarly journals Determination of the Domain of the Lactobacillus delbrueckii subsp. bulgaricus Cell Surface Proteinase PrtB Involved in Attachment to the Cell Wall after Heterologous Expression of the prtB Gene in Lactococcus lactis

2003 ◽  
Vol 69 (6) ◽  
pp. 3377-3384 ◽  
Author(s):  
Jacques-Edouard Germond ◽  
Mich�le Delley ◽  
Christophe Gilbert ◽  
Dani�le Atlan
Keyword(s):  
Amino Acids ◽  
Cell Wall ◽  
Cell Surface ◽  
Lactococcus Lactis ◽  
Culture Medium ◽  
Surface Display ◽  
Cell Surface Display ◽  
Lactobacillus Delbrueckii ◽  
Heterologous Proteins

ABSTRACT Belonging to the subtilase family, the cell surface proteinase (CSP) PrtB of Lactobacillus delbrueckii subsp. bulgaricus differs from other CSPs synthesized by lactic acid bacteria. Expression of the prtB gene under its own promoter was shown to complement the proteinase-deficient strain MG1363 (PrtP− PrtM−) of Lactococcus lactis subsp. cremoris. Surprisingly, the maturation process of PrtB, unlike that of lactococcal CSP PrtPs, does not require a specific PrtM-like chaperone. The carboxy end of PrtB was previously shown to be different from the consensus anchoring region of other CSPs and exhibits an imperfect duplication of 59 amino acids with a high lysine content. By using a deletion strategy, the removal of the last 99 amino acids, including the degenerated anchoring signal (LPKKT), was found to be sufficient to release a part of the truncated PrtB into the culture medium and led to an increase in PrtB activity. This truncated PrtB is still active and enables L. lactis MG1363 to grow in milk supplemented with glucose. By contrast, deletion of the last 806 amino acids of PrtB led to the secretion of an inactive proteinase. Thus, the utmost carboxy end of PrtB is involved in attachment to the bacterial cell wall. Proteinase PrtB constitutes a powerful tool for cell surface display of heterologous proteins like antigens.

Construction of a Yeast Cell-Surface Display System and Expression of Trametes sp. laccase

2011 ◽  
Vol 347-353 ◽  
pp. 3635-3640 ◽  
Author(s):  
Jian Zhang Lu ◽  
Qin Guo ◽  
Mei Lin Cui ◽  
Lu Yang ◽  
Shan Shan Du ◽  
...  
Keyword(s):  
Cell Surface ◽  
Surface Display ◽  
Cell Surface Display ◽  
Heterologous Proteins ◽  
Secretion Signal ◽  
Yeast Cell Surface ◽  
Gal1 Promoter ◽  
Genes Encoding ◽  
Display Technology ◽  
A Cell

Laccases (1.10.3.2, p-diphenol: dioxygen oxidoreductases) is a family of blue copper-containing oxidases that are commonly found in bacteria, fungi and plants. It is able to oxidize and degrade a variety of aromatic compounds and other organic compounds. Due to this ability, laccases can serve environmental bioremediation processes and industrial purposes. Cell-surface display of enzymes is one of the most attractive applications in yeast. It is a effective utilization to construct the whole cell biocatalyst. The cDNA sequence of Trametes sp. C30 LAC3 was optimized and synthesized according to the codon bias of Saccharomyces Italic textcerevisiae, because codon optimization has been proved to be effective to maximize production of heterologous proteins in yeast. The genes encoding galactokinase (GAL1) promoter, α-mating factor 1 (MFα1) pre-pro secretion signal, fully codon-optimized LAC3, the 320 amino acids of C terminal of α-agglutinin, alcohol dehydrogenase (ADH1) terminator and kanMX cassette were amplified and cloned into YEplac181 to construct a cell-surface display vector called pGMAAK-lac3 with α-agglutinin as an anchor. Then pGMAAK-lac3 was transformed into S. cerevisiae. The results show LAC3 was immobilized and actively expressed on S. cerevisiae. However, the substrate specifity and activity were obviously changed. The displayed LAC3 lost the activity to phenolic substrate (guaiacol) and its activity to non-phenolic substrate (ABTS) was greatly reduced. To our knowledge, this was the first attempt to construct and express laccase through cell-surface display technology.

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