Alp, a suppressor of lon protease mutants in Escherichia coli.

Foodborne pathogens are a public health threat globally. Shiga toxin-producing Escherichia coli (STEC), particularly O26, O111, and O157 STEC, are often associated with foodborne illness in humans. To create effective preharvest interventions, it is critical to understand which factors STEC strains use to colonize the gastrointestinal tract of cattle, which serves as the reservoir for these pathogens. Several colonization factors are known, but little is understood about initial STEC colonization factors. Our objective was to identify these factors via contrasting gene expression between nonpathogenic E. coli and STEC. Colonic explants were inoculated with nonpathogenic E. coli strain MG1655 or STEC strains (O26, O111, or O157), bacterial colonization levels were determined, and RNA was isolated and sequenced. STEC strains adhered to colonic explants at numerically but not significantly higher levels compared to MG1655. After incubation with colonic explants, flagellin (fliC) was upregulated (log2 fold-change = 4.0, p < 0.0001) in O157 STEC, and collectively, Lon protease (lon) was upregulated (log2 fold-change = 3.6, p = 0.0009) in STEC strains compared to MG1655. These results demonstrate that H7 flagellum and Lon protease may play roles in early colonization and could be potential targets to reduce colonization in cattle.

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Front Cover: An Integrated Proteomic Approach Uncovers Novel Substrates and Functions of the Lon Protease in Escherichia coli

PROTEOMICS ◽

10.1002/pmic.201870111 ◽

2018 ◽

Vol 18 (13) ◽

pp. 1870111

Author(s):

Jan Arends ◽

Marcena Griego ◽

Nikolas Thomanek ◽

Claudia Lindemann ◽

Blanka Kutscher ◽

...

Keyword(s):

Escherichia Coli ◽

Lon Protease ◽

Front Cover ◽

Proteomic Approach

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Saturation and specificity of the Lon protease of Escherichia coli.

Journal of Bacteriology ◽

10.1128/jb.172.12.7098-7103.1990 ◽

1990 ◽

Vol 172 (12) ◽

pp. 7098-7103 ◽

Cited By ~ 16

Author(s):

E Dervyn ◽

D Canceill ◽

O Huisman

Keyword(s):

Escherichia Coli ◽

Lon Protease

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Regulation of RcsA by the ClpYQ (HslUV) protease in Escherichia coli

Microbiology ◽

10.1099/mic.0.26446-0 ◽

2004 ◽

Vol 150 (2) ◽

pp. 437-446 ◽

Cited By ~ 23

Author(s):

Mei-Shiue Kuo ◽

Kuei-Peng Chen ◽

Whi Fin Wu

Keyword(s):

Escherichia Coli ◽

Lon Protease ◽

Specific Substrate ◽

Galactosidase Activity ◽

Wild Type ◽

Reporter Construct ◽

Fusion Construct ◽

In The Wild ◽

A Cell ◽

Cell Division Inhibitor

Escherichia coli ClpYQ protease and Lon protease possess a redundant function for degradation of SulA, a cell division inhibitor. An experimental cue implied that the capsule synthesis activator RcsA, a known substrate of Lon, is probably a specific substrate for the ClpYQ protease. This paper shows that overexpression of ClpQ and ClpY suppresses the mucoid phenotype of a lon mutant. Since the cpsB (wcaB) gene, involved in capsule synthesis, is activated by RcsA, the reporter construct cpsB–lacZ was used to assay for β-galactosidase activity and thus follow RcsA stability. The expression of cpsB–lacZ was increased in double mutants of lon in combination with clpQ or/and clpY mutation(s) compared with the wild-type or lon single mutants. Overproduction of ClpYQ or ClpQ decreased cpsB–lacZ expression. Additionally, a PBAD–rcsA fusion construct showed quantitatively that an inducible RcsA activates cpsB–lacZ expression. The effect of RcsA on cpsB–lacZ expression was shown to be influenced by the ClpYQ activities. Moreover, a rcsA Red –lacZ translational fusion construct showed higher activity of RcsARed–LacZ in a clpQ clpY strain than in the wild-type. By contrast, overproduction of cellular ClpYQ resulted in decreased β-galactosidase levels of RcsARed–LacZ. Taken together, the data indicate that ClpYQ acts as a secondary protease in degrading the Lon substrate RcsA.

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Heteromeric Interactions among Nucleoid-Associated Bacterial Proteins: Localization of StpA-Stabilizing Regions in H-NS of Escherichia coli

Journal of Bacteriology ◽

10.1128/jb.183.7.2343-2347.2001 ◽

2001 ◽

Vol 183 (7) ◽

pp. 2343-2347 ◽

Cited By ~ 50

Author(s):

Jörgen Johansson ◽

Sven Eriksson ◽

Berit Sondén ◽

Sun Nyunt Wai ◽

Bernt Eric Uhlin

Keyword(s):

Escherichia Coli ◽

Stable Form ◽

Lon Protease ◽

Wild Type ◽

Bacterial Proteins ◽

Regulatory Systems ◽

Associated Proteins ◽

Gene Regulatory

ABSTRACT The nucleoid-associated proteins H-NS and StpA inEscherichia coli bind DNA as oligomers and are implicated in gene regulatory systems. There is evidence for both homomeric and heteromeric H-NS–StpA complexes. The two proteins show differential turnover, and StpA was previously found to be subject to protease-mediated degradation by the Lon protease. We investigated which regions of the H-NS protein are able to prevent degradation of StpA. A set of truncated H-NS derivatives was tested for their ability to mediate StpA stability and to form heteromers in vitro. The data indicate that H-NS interacts with StpA at two regions and that the presence of at least one of the H-NS regions is necessary for StpA stability. Our results also suggest that a proteolytically stable form of StpA, StpAF21C, forms dimers, whereas wild-type StpA in the absence of H-NS predominantly forms tetramers or oligomers, which are more susceptible to proteolysis.

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Production of Recombinant Proteins in the lon-Deficient BL21(DE3) Strain of Escherichia coli in the Absence of the DnaK Chaperone

Applied and Environmental Microbiology ◽

10.1128/aem.00255-09 ◽

2009 ◽

Vol 75 (11) ◽

pp. 3803-3807 ◽

Cited By ~ 17

Author(s):

Julien Ratelade ◽

Marie-Caroline Miot ◽

Emmett Johnson ◽

Jean-Michel Betton ◽

Philippe Mazodier ◽

...

Keyword(s):

Escherichia Coli ◽

Recombinant Proteins ◽

Protein Production ◽

Lon Protease ◽

Unfolded Proteins ◽

Unique Approach

ABSTRACT To eliminate unavoidable contamination of purified recombinant proteins by DnaK, we present a unique approach employing a BL21(DE3) ΔdnaK strain of Escherichia coli. Selected representative purified proteins remained soluble, correctly assembled, and active. This finding establishes DnaK dispensability for protein production in BL21(DE3), which is void of Lon protease, key to eliminating unfolded proteins.

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Regulated Expression of the Escherichia coli dam Gene

Journal of Bacteriology ◽

10.1128/jb.185.16.5012-5014.2003 ◽

2003 ◽

Vol 185 (16) ◽

pp. 5012-5014 ◽

Cited By ~ 13

Author(s):

Melissa A. Calmann ◽

M. G. Marinus

Keyword(s):

Escherichia Coli ◽

Binding Site ◽

Ribosome Binding Site ◽

Lon Protease ◽

Ribosome Binding ◽

Regulated Expression

ABSTRACT Regulated expression of the Escherichia coli dam gene has been achieved with the araBAD promoter lacking a ribosome binding site. Cultures of dam mutants containing plasmid pMQ430 show no detectable methylation in the absence of arabinose and complete methylation in its presence. Dam methyltransferase is a substrate for the Lon protease.

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