Autolyse the cell in order to save it? Inducing, then blocking, autolysis as a strategy for delaying cell death in the probiotic Lactobacillus reuteri

Zimmerman, Tahl; Gyawali, Rabin; Ibrahim, Salam A.

doi:10.1007/s10529-017-2380-8

Cited by 8 publications

(14 citation statements)

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“…Importantly, we did not observe the suppression of downstream lipoteichoic acid production that is the expected consequence of choline kinase inhibition 17 due to the putative attenuation of phosphocholine production which would remove one of the building blocks of LTA Instead, we observed increased phosphocholine production (in the case of MN58b) and changes in lipoteichic acid sizes to either smaller (MN58b) or larger sizes (RSM-932A). This change appears to have an effect on cell sensitivity to autolysis and on the shape of the cell wall.…”

Section: Discussioncontrasting

confidence: 53%

“…Although autolysin proteins would be released, they would not be able to dock on the cell surface because the autolysin choline binding domains would have fewer choline containing LTA molecules with which to interact. Both LTA reduction and cell sensitization had previously been observed with HC-3 17 . Surprisingly, cells grown in the presence of 0.5 MIC RSM-932A were equally sensitive to autolysis as untreated controls, while MN58b actually primed the cells to be autolyzed (Fig.…”

Section: Resultsmentioning

confidence: 64%

“…Autolysis is a process of cell lysis and death that is mediated by LTAs via choline binding autolysins 19 such as LytA. The choline analog and sChoK inhibitor HC-3 had previously been found to attenuate the autolytic process in Lactobacillus reuteri cells 17 . In light of the non-specificity observed with HC-3 (being able to interfere with both ChoKs and the autolytic process), it seemed logical to check the specificity of MN58b and RSM-932A for sChoK inhibition.…”

Section: Resultsmentioning

confidence: 99%

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Identification and validation of novel and more effective choline kinase inhibitors against Streptococcus pneumoniae

Zimmerman

Chasten

Lacal

et al. 2020

Sci Rep

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Streptococcus pneumoniae choline kinase (sChoK) has previously been proposed as a drug target, yet the effectiveness of the first and only known inhibitor of sChoK, HC-3, is in the millimolar range. The aim of this study was thus to further validate sChoK as a potential therapeutic target by discovering more powerful sChoK inhibitors. LDH/PK and colorimetric enzymatic assays revealed two promising sChoK inhibitor leads RSM-932A and MN58b that were discovered with IC50 of 0.5 and 150 μM, respectively, and were shown to be 2–4 magnitudes more potent than the previously discovered inhibitor HC-3. Culture assays showed that the minimum inhibitory concentration (MIC) of RSM-932A and MN58b for S. pneumoniae was 0.4 μM and 10 μM, respectively, and the minimum lethal concentration (MLC) was 1.6 μM and 20 μM, respectively. Western blot monitoring of teichoic acid production revealed differential patterns in response to each inhibitor. In addition, both inhibitors possessed a bacteriostatic mechanism of action, and neither interfered with the autolytic effects of vancomycin. Cells treated with MN58b but not RSM-932A were more sensitive to a phosphate induced autolysis with respect to the untreated cells. SEM studies revealed that MN58b distorted the cell wall, a result consistent with the apparent teichoic acid changes. Two novel and more highly potent putative inhibitors of sChoK, MN58b and RSM-932A, were characterized in this study. However, the effects of sChoK inhibitors can vary at the cellular level. sChoK inhibition is a promising avenue to follow in the development of therapeutics for treatment of S. pneumoniae.

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Section: Discussioncontrasting

confidence: 53%

Section: Resultsmentioning

confidence: 64%

Section: Resultsmentioning

confidence: 99%

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Identification and validation of novel and more effective choline kinase inhibitors against Streptococcus pneumoniae

Zimmerman

Chasten

Lacal

et al. 2020

Sci Rep

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“…If the production of lipoteichoic acid was reduced on the cell wall as a consequence of treatment with HC-3, one would expect cells treated with HC-3 to be more resistant to autolysis. Lipoteichoic acid is the component of the cell wall that anchors the autolytic protein autolysin in the first step of a two-step process which leads to autolysin cleavage of peptidoglycan and subsequent cell lysis [ 19 ]. By reducing the production of lipoteichoic acid, fewer molecules of autolysin can bind to the cell surface.…”

Section: Resultsmentioning

confidence: 99%

“…To test this idea, control and treated cells were exposed to a deoxycholate/phosphate solution. Phosphate is known to induce autolysis in Gram-positive bacterial cells [ 19 ], and deoxycholate, a detergent, has been known to induce the release of autolysin into solution, allowing the autolysin to interact with the cell wall, leading to cell lysis [ 20 ]. Cells grown in the presence of HC-3 were more resistant to autolysis induced by this detergent mixture with respect to a non-autolytic control (see Figure 3 B).…”

Section: Resultsmentioning

confidence: 99%

Choline Kinase, A Novel Drug Target for the Inhibition of Streptococcus pneumoniae

Zimmerman

Ibrahim

2017

Antibiotics

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Gram-positive pathogens, such as Streptococcus pneumoniae, can have deleterious effects on both human and animal health. Antibiotics and antimicrobials have been developed to treat infections caused by such pathogens and to prevent food contamination. However, these strategies have been increasingly thwarted by the emergence of resistant bacteria strains. Thus, new methods for controlling Gram-positive pathogen growth need to be continuously developed. Choline analogs, such as Hemicholinium-3 (HC-3), have been shown to be useful in blocking cell division in eukaryotic cells through the inhibition of choline kinase, an enzyme which catalyzes the production of phosphocholine from choline and ATP. In some Gram-positive pathogens, choline kinase is an important enzyme in the production of the cell wall element, lipoteichoic acid. However, it is not known if inhibiting this enzyme has any effect on cell division in Gram-positive bacteria. Using the R6 strain as a model, we tested the ability of HC-3 to block the activity of choline kinase in S. pneumoniae and inhibit cell growth. Mass-spectrometry measurements of crude extracts revealed that HC-3 blocked choline kinase activity. Turbidity measurements and population counts showed that HC-3 inhibited cell growth. Competition assays with choline suggested that HC-3 also blocked choline transporters. Western blots showed that lipoteichoic acid production was blocked in the presence of HC-3, and autolytic assays showed that this decrease in lipoteichoic acids caused cells to be more resistant to autolysis. Scanning electron microscopy revealed that HC-3 distorted the cell wall. This study thus establishes choline kinase as a novel drug target for S. pneumoniae.

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Topical Delivery of Lactobacillus Culture Supernatant Increases Survival and Wound Resolution in Traumatic Acinetobacter baumannii Infections

Stanbro

Park

Bond

et al. 2019

Probiotics & Antimicro. Prot.

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Autolyse the cell in order to save it? Inducing, then blocking, autolysis as a strategy for delaying cell death in the probiotic Lactobacillus reuteri

Abstract: Inducing autolysis and then blocking it with choline and its analogs is a promising approach for retaining the viability of L. reuteri cells.

Cited by 8 publications

References 9 publications

Identification and validation of novel and more effective choline kinase inhibitors against Streptococcus pneumoniae

Identification and validation of novel and more effective choline kinase inhibitors against Streptococcus pneumoniae

Choline Kinase, A Novel Drug Target for the Inhibition of Streptococcus pneumoniae

Topical Delivery of Lactobacillus Culture Supernatant Increases Survival and Wound Resolution in Traumatic Acinetobacter baumannii Infections

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