Generation of catalytic protein particles in Escherichia coli cells using the cellulose-binding domain from Cellulomonas fimi as a fusion partner

2011 ◽  
Vol 16 (6) ◽  
pp. 1173-1179 ◽  
Author(s):  
Su-Lim Choi ◽  
Sang Jun Lee ◽  
Jae-Seok Ha ◽  
Jae Jun Song ◽  
Young Ha Rhee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Binding Domain ◽  
Fusion Partner ◽  
Cellulose Binding Domain ◽  
Cellulomonas Fimi ◽  
Escherichia Coli Cells ◽  
Protein Particles ◽  
Cellulose Binding

scholarly journals Enzyme Immobilization Using the Cellulose-Binding Domain of a Cellulomonas Fimi Exoglucanase

1989 ◽  
Vol 7 (6) ◽  
pp. 604-607 ◽  
Author(s):  
Edgar Ong ◽  
Neil R. Gilkes ◽  
R. Antony J. Warren ◽  
Robert C. Miller ◽  
Douglas G. Kilburn
Keyword(s):  
Enzyme Immobilization ◽  
Binding Domain ◽  
Cellulose Binding Domain ◽  
Cellulomonas Fimi ◽  
Cellulose Binding

The cellulose-binding domain of endoglucanase A (CenA) from Cellulomonas fimi: evidence for the involvement of tryptophan residues in binding

1994 ◽  
Vol 11 (4) ◽  
pp. 747-755 ◽  
Author(s):  
Neena Din ◽  
Ian J. Forsythe ◽  
Leslie D. Burtnick ◽  
Neil R. Gilkes ◽  
Robert C. Miller ◽  
...  
Keyword(s):  
Binding Domain ◽  
Cellulose Binding Domain ◽  
Cellulomonas Fimi ◽  
Tryptophan Residues ◽  
Cellulose Binding

scholarly journals Specific Adhesion to Cellulose and Hydrolysis of Organophosphate Nerve Agents by a Genetically Engineered Escherichia coli Strain with a Surface-Expressed Cellulose-Binding Domain and Organophosphorus Hydrolase

2002 ◽  
Vol 68 (4) ◽  
pp. 1684-1689 ◽  
Author(s):  
Aijun A. Wang ◽  
Ashok Mulchandani ◽  
Wilfred Chen
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Nerve Agents ◽  
Genetically Engineered ◽  
Binding Domain ◽  
Organophosphorus Hydrolase ◽  
Fusion System ◽  
Cellulose Binding Domain ◽  
Cellulose Binding ◽  
Hydrolysis Of

ABSTRACT A genetically engineered Escherichia coli cell expressing both organophosphorus hydrolase (OPH) and a cellulose-binding domain (CBD) on the cell surface was constructed, enabling the simultaneous hydrolysis of organophosphate nerve agents and immobilization via specific adsorption to cellulose. OPH was displayed on the cell surface by use of the truncated ice nucleation protein (INPNC) fusion system, while the CBD was surface anchored by the Lpp-OmpA fusion system. Production of both INPNC-OPH and Lpp-OmpA-CBD fusion proteins was verified by immunoblotting, and the surface localization of OPH and the CBD was confirmed by immunofluorescence microscopy. Whole-cell immobilization with the surface-anchored CBD was very specific, forming essentially a monolayer of cells on different supports, as shown by electron micrographs. Optimal levels of OPH activity and binding affinity to cellulose supports were achieved by investigating expression under different induction levels. Immobilized cells degraded paraoxon rapidly at an initial rate of 0.65 mM/min/g of cells (dry weight) and retained almost 100% efficiency over a period of 45 days. Owing to its superior degradation capacity and affinity to cellulose, this immobilized-cell system should be an attractive alternative for large-scale detoxification of organophosphate nerve agents.

Enzyme-linked assay of cellulose-binding domain functions from Cellulomonas fimi on multi-well microtiter plate

2013 ◽  
Vol 18 (3) ◽  
pp. 575-580 ◽  
Author(s):  
Hyeon-Dong Kim ◽  
Su-Lim Choi ◽  
Haseong Kim ◽  
Jung Hoon Sohn ◽  
Seung-Goo Lee
Keyword(s):  
Microtiter Plate ◽  
Binding Domain ◽  
Cellulose Binding Domain ◽  
Cellulomonas Fimi ◽  
Cellulose Binding

scholarly journals Spontaneous Cleavages of a Heterologous Protein, the CenA Endoglucanase of Cellulomonas fimi, in Escherichia coli

2021 ◽  
Vol 14 ◽  
pp. 117863612110246
Author(s):  
Cheuk Yin Lai ◽  
Ka Lun Ng ◽  
Hao Wang ◽  
Chui Chi Lam ◽  
Wan Keung Raymond Wong
Keyword(s):  
Escherichia Coli ◽  
Cellulolytic Bacterium ◽  
Systematic Screening ◽  
Cellulomonas Fimi ◽  
E Coli ◽  
Substrate Complex ◽  
Enzyme Substrate ◽  
Glycosylation Sites ◽  
Endogenous Proteins ◽  
Cellulose Binding

CenA is an endoglucanase secreted by the Gram-positive cellulolytic bacterium, Cellulomonas fimi, to the environment as a glycosylated protein. The role of glycosylation in CenA is unclear. However, it seems not crucial for functional activity and secretion since the unglycosylated counterpart, recombinant CenA (rCenA), is both bioactive and secretable in Escherichia coli. Using a systematic screening approach, we have demonstrated that rCenA is subjected to spontaneous cleavages (SC) in both the cytoplasm and culture medium of E. coli, under the influence of different environmental factors. The cleavages were found to occur in both the cellulose-binding (CellBD) and catalytic domains, with a notably higher occurring rate detected in the former than the latter. In CellBD, the cleavages were shown to occur close to potential N-linked glycosylation sites, suggesting that these sites might serve as ‘attributive tags’ for differentiating rCenA from endogenous proteins and the points of initiation of SC. It is hypothesized that glycosylation plays a crucial role in protecting CenA from SC when interacting with cellulose in the environment. Subsequent to hydrolysis, SC would ensure the dissociation of CenA from the enzyme-substrate complex. Thus, our findings may help elucidate the mechanisms of protein turnover and enzymatic cellulolysis.

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