A Mechanism-Based Whole-Cell Screening Assay to Identify Inhibitors of Protein Export in Escherichia coli by the Sec Pathway

Crowther, Gregory J.; Quadri, Shafat A.; Shannon-Alferes, Benjamin J.; Voorhis, Wesley C. Van; Rosen, Henry

doi:10.1177/1087057111431606

Cited by 11 publications

(11 citation statements)

References 17 publications

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“…The IL-6 levels in the study group were lower than in the control group on the tenth day after the operation (P < 0.05; Table 4). stages after surgery (Bonekamp et al, 2010;Tannu et al, 2010;Crowther et al, 2012). In this study, gastric cancer patients received EN support with different nutritional preparations during different perioperative periods, which were chosen depending on functional conditions of the digestive tract (Ichimaru et al, 2012).…”

Section: Comparison Of Changes In Inflammatory Reaction Indicators Bementioning

confidence: 99%

Impact of enteral nutrition on postoperative immune function and nutritional status

Wang¹,

Hou²,

Wu³

et al. 2015

Genet. Mol. Res.

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ABSTRACT. We studied the effects of enteral nutrition (EN) support initiated 1 week before surgery on postoperative nutritional status, immune function, and inflammatory response in gastric cancer patients. A total of 200 gastric cancer patients were randomly divided into two groups: EN starting 1 week before surgery (study group) and EN starting early after surgery (control group). The two groups received EN support, following different therapeutic schedules, until the 9th day after operation. In the patients, body weight, skinfold thickness, upper-arm circumference, white blood cell count, albumin, prealbumin, C-reactive protein, peripheral immunoglobulins (IgA, IgG, and IgM), T lymphocyte subsets, interleukin-6, and tumor necrosis factor-α were measured 10 days before and after surgery and on the first day after surgery. There was no statistically significant difference in the results of recovery time of passage of gas by anus, abdominal distension, stomachache, blood glucose, hepatic and renal functions, and electrolytes between the two groups of patients (P > 0. 05). Adverse reactions occurred to both groups at 1 and 2 days after operation. Such conditions was improved after the intravenous drip rate was adjusted. The albumin and prealbumin levels of the patients in both groups decreased at 1 day after operation (P < 0. 05). The levels rose when the research was finished (P < 0. 05). The prealbumin level of the study group was higher than that of the control group at 10 days after operation (P < 0. 05). The IgG level of the study group was higher than that of the control group at 10 days after operation (P < 0. 05). The two groups of inflammatory reaction indicators of the study group were lower than those of the control group at 10 days after operation (P < 0. 05). This study indicates that appropriate preoperative EN support for gastric cancer patients can improve their postoperative nutritional status and immune function, can reduce inflammatory response, and is more conducive to the recovery of patients.

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Section: Comparison Of Changes In Inflammatory Reaction Indicators Bementioning

confidence: 99%

Impact of enteral nutrition on postoperative immune function and nutritional status

Wang¹,

Hou²,

Wu³

et al. 2015

Genet. Mol. Res.

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“…Crowther et al [153] have developed a whole-cell-based inhibitor screening strategy for the SecA machinery and demonstrated its high-throughput utility using various inhibitors. It involves using an engineered E. coli strain, which produces beta-galactosidase protein (β-gal) having the LamB sequence, causing it to be transported via Sec machinery.…”

Section: Reported Seca Inhibitorsmentioning

confidence: 99%

SecA: A Potential Antimicrobial Target

et al. 2015

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There is a consensus in the medical profession of the pressing need for novel antimicrobial agents due to issues related to drug resistance. In practice, solutions to this problem to a large degree lie with the identification of new and vital targets in bacteria and subsequently designing their inhibitors. We consider SecA a very promising antimicrobial target. In this review, we compile and analyze information available on SecA to show that inhibition of SecA has a multitude of consequences. Furthermore, we discuss issues critical to the design and evaluation of SecA inhibitors.

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“…Our protocol was essentially as described previously 8 aside from adaptations to the high-throughput screening environment. In brief, EC626 cells suspended in 3 mL of Miller's Lysogeny broth (LB), with 0.2% glucose (w/v) added to suppress LacZ expression, were grown overnight in 15 mL round-bottom tubes at 37 °C under vigorous aeration.…”

Section: High-throughput Screenmentioning

confidence: 99%

“…We have taken a more inclusive approach to Sec pathway drug development by developing a screening assay that should identify inhibitors of protein export by the Sec pathway as a whole, rather than inhibitors of SecA alone. 8 Our assay uses an Escherichia coli strain (EC626) that expresses a recombinant beta-galactosidase (β-gal) with a signal peptide that sends it through the SecYEG channel to the periplasm, where the oxidizing environment does not allow conformational maturation to an active enzyme. Thus, any β-gal activity seen in these cells is cytoplasmic and indicates an impairment of export through SecYEG.…”

Section: Introductionmentioning

confidence: 99%

The Bacterial Sec Pathway of Protein Export: Screening and Follow-Up

et al. 2015

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Most noncytoplasmic bacterial proteins are exported through the SecYEG channel in the cytoplasmic membrane. This channel and its associated proteins, collectively referred to as the Sec pathway, have strong appeal as a possible antibiotic drug target, yet progress toward new drugs targeting this pathway has been slow, perhaps due partly to many researchers' focus on a single component, the SecA ATPase. Here we report on a pathway-based screen in which beta-galactosidase (β-gal) activity is trapped in the cytoplasm of Escherichia coli cells if translocation through SecYEG is impaired. Several hit compounds passed a counterscreen distinguishing between β-gal overexpression and impaired β-gal export. However, the most extensively characterized hit gave limited E. coli growth inhibition (EC(50) ≥ 400 µM), and growth inhibition could not be unambiguously linked to the compound's effect on the Sec pathway. Our study and others underscore the challenges of finding potent druglike hits against this otherwise promising drug target.

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A Mechanism-Based Whole-Cell Screening Assay to Identify Inhibitors of Protein Export in Escherichia coli by the Sec Pathway

Cited by 11 publications

References 17 publications

Impact of enteral nutrition on postoperative immune function and nutritional status

Impact of enteral nutrition on postoperative immune function and nutritional status

SecA: A Potential Antimicrobial Target

The Bacterial Sec Pathway of Protein Export: Screening and Follow-Up

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