• Biomedical Science Letters
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• v.5 no.1
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• pp.27-40
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• 1999

This study was focused on the evaluation of MarB alongside with MarA for its regulatory effects upon the efflux function of AcrAB pump, which were induced or not, perhaps as a target. Transductions of marR and/or acrAB mutation which were derived from Mar and/or AcrAB mutants of wild type E. coli K-12, respectively, into the multicopy plasmid in wild type E. coli backgrounds or into the chromosome of isogenic parents were done. Minimal inhibitory concentration (MIC) of transduced mutants was compared with their original mutants. This study reports the indirect evidences that suggests a model in which MarB along with MarA have a putative negative regulatory effect upon the efflux function of AcrAB/TolC pump while MarA alone have a positive regulatory effect to the expression of acrRAB operon at transcription level. The target of MarB with MarA for its putative negative regulator might be the AcrAB efflux pump. Another efflux system (s) might be negatively regulated by MarB with MarA, and be involved in the efflux of antibiotics which were otherwise extruded preferentially by AcrAB efflux pump.

• Biomedical Science Letters
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• v.27 no.2
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• pp.59-68
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• 2021

Multidrug-resistant strain of Acinetobacter baumannii (MDRAB) is an emerging pathogen in health care facilities, preventing MDRAB is a public health concern. We conducted this experiment on a clinical isolate of A. baumannii with two main goals: the role of the efflux pump system in the stress provision of carbapenem and the response to the transcription level of the efflux pump gene. A total of 34 strains of A. baumannii was isolated from the Yangsan Hospital of Pusan National University. First, when we compared and observed the expression of the efflux pump gene and antibacterial resistance to carbapenem, a strong correlation was observed between carbapenem resistance and overexpression of adeB (P=0.0056). Second, a correlation between the efflux pump and concentration gradient and tolerance to carbapenem stress at the AdeABC efflux pump genes transcription level was confirmed. Our results revealed that the expression of the AdeABC efflux pump is an important resistance determinant in obtaining antibiotic resistance of the carbapenem group in A. baumannii.

• Microbiology and Biotechnology Letters
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• v.31 no.3
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• pp.230-234
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• 2003

The Bacillus subtilis YvaE protein, the small multidrug resistance (SMR) family (TC #2.A. 7.1), is shown to catalyze efflux of multiple cationic drugs including many disinfectants, when it was cloned and expressed in Escherichia coli. When the yvaD gene was coexpressed with yvaE gene, the yvaD protein, encoded within a single operon with the yvaE gene, is shown to counteract the action ofYvaE. By ethidium efflux analysis, the cells harvoring a vector with yvaE gene showed a rapid ethidium efflux, compared with the control cells. These results clearly suggest that YvaE mediates drug export from the cell cytoplasm.

• Journal of Microbiology and Biotechnology
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• v.23 no.3
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• pp.430-435
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• 2013

Multidrug resistance, especially multidrug efflux mechanisms that extrude structurally unrelated cytotoxic compounds from the cell by multidrug transporters, is a serious problem and one of the main reasons for the failure of therapeutic treatment of infections by pathogenic microorganisms as well as of cancer cells. Streptococcus mutans is considered one of the primary causative agents of dental caries and periodontal disease, which comprise the most common oral diseases. A fragment of chromosomal DNA from S. mutans KCTC3065 was cloned using Escherichia coli KAM32 as host cells lacking major multidrug efflux pumps. Although E. coli KAM32 cells were very sensitive to many antimicrobial agents, the transformed cells harboring a recombinant plasmid became resistant to several structurally unrelated antimicrobial agents such as tetracycline, kanamycin, rhodamin 6G, ampicillin, acriflavine, ethidium bromide, and tetraphenylphosphonium chloride. This suggested that the cloned DNA fragment carries a gene encoding a multidrug efflux pump. Among 49 of the multidrug-resistant transformants, we report the functional gene cloning and characterization of the function of one multidrug efflux pump, namely MdeA from S. mutans, which was expressed in E. coli KAM32. Judging from the structural and biochemical properties, we concluded that MdeA is the first cloned and characterized multidrug efflux pump using the proton motive force as the energy for efflux drugs.

• Journal of Microbiology
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• v.39 no.1
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• pp.62-66
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• 2001

Unlike eukaryotic efflux pumps energized by ATPase bacterial efflux pumps are energized by the proton motive force. That is the reason why CCCP, an inhibitor of proton motive forcer is widely used to study the bacterial efflux pump. In many cases, efflux systems have been observed only in quinolone-resistant bacteria. Most of the quinolone-susceptible strains have been found to maintain little efflux pump. However some susceptible bacteria skewed the increased intracellular quinolone concentration only at a low concentration (0.01 or 0.1 mM) but net at a high concentration (1 mM) of CCCP. If bacterial cells were killed at high concentrations of CCCP and lost the integrity of their membranes, the intracellular quinolone would leak out from cells with no efflux system. The efflux pump system in the quinolone-susceptible strains could net be detected at the same concentration used for resistant bacteria. To test this hypothesist the intracellular quinolone concentration in the quinolone-susceptible and -resistant strains of Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus was assayed at various concentrations of CCCP. Since the effect of CCCP is very rapid, the survival of bacteria was observed by assaying the DNA synthesis in 5 min. In the case of E. coli, but not P. aeruginosa or S. aureus, the quinolone susceptible strain was more susceptible to CCCP than the quinolone resistant ones, especially when the incubation with CCCP was extended. Decrease of the intracellular quinolone concentration resulted in a false result-no or weak efflux system in the quinolone susceptible strains. Results suggested that the concentration of CCCP should be optimized in order to detect the efflux system in the quinolone susceptible strains of E. coli.

• BMB Reports
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• v.56 no.6
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• pp.326-334
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• 2023

Antibiotic resistance (AR) is a silent pandemic that kills millions worldwide. Although the development of new therapeutic agents against antibiotic resistance is in urgent demand, this has presented a great challenge, especially for Gram-negative bacteria that have inherent drug-resistance mediated by impermeable outer membranes and multidrug efflux pumps that actively extrude various drugs from the bacteria. For the last two decades, multidrug efflux pumps, including AcrAB-TolC, the most clinically important efflux pump in Gram-negative bacteria, have drawn great attention as strategic targets for re-sensitizing bacteria to the existing antibiotics. This article aims to provide a concise overview of the AcrAB-TolC operational mechanism, reviewing its architecture and substrate specificity, as well as the recent development of AcrAB-TolC inhibitors.

• Biomolecules & Therapeutics
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• v.14 no.1
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• pp.45-49
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• 2006

The purpose of this study was to clarify the efflux transport system of choline from brain to blood across the blood-brain barrier (BBB) in rats using the brain efflux index (BEI) method. $[^3H]$Choline was micro-injected into parietal cortex area 2 (Par2) of the rat brain, and was eliminated from the brain with elimination halflife of 45 min. The BBB efflux clearance of $[^3H]$choline was about 124 mL/min/g brain, which was determined from combination of an elimination rate constant $(1.54X10^{-2}min^{-1})$ and the distribution volume in the brain (8.05 mL/g brain). The efflux of $[^3H]$choline was inhibited by unlabeled choline in a dose-dependent manner and was significantly inhibited by cationic substrates, such as hemicholinium-3 and tetraethylammonium (TEA). These results suggest that the BBB may act as an efflux pump for choline to reduce the excessive choline concentration in the brain interstitial fluid.

• Journal of Microbiology and Biotechnology
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• v.30 no.7
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• pp.982-995
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• 2020

A putative multidrug efflux gene, yddA, was cloned from the Escherichia coli K-12 strain. A drug-sensitive strain of E. coli missing the main multidrug efflux pump AcrB was constructed as a host and the yddA gene was knocked out in wild-type (WT) and drug-sensitive E. coliΔacrB to study the yddA function. Sensitivity to different substrates of WT E.coli, E. coliΔyddA, E. coliΔacrB and E. coliΔacrBΔyddA strains was compared with minimal inhibitory concentration (MIC) assays and fluorescence tests. MIC assay and fluorescence test results showed that YddA protein was a multidrug efflux pump that exported multiple substrates. Three inhibitors, ortho-vanadate, carbonyl cyanide m-chlorophenylhydrazone (CCCP), and reserpine, were used in fluorescence tests. Ortho-vanadate and reserpine significantly inhibited the efflux and increased accumulation of ethidium bromide and norfloxacin, while CCCP had no significant effect on YddA-regulated efflux. The results indicated that YddA relies on energy released from ATP hydrolysis to transfer the substrates and YddA is an ABC-type multidrug exporter. Functional study of unknown ATP-binding cassette (ABC) superfamily transporters in the model organism E. coli is conducive to discovering new multidrug resistance-reversal targets and providing references for studying other ABC proteins of unknown function.

• BMB Reports
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• v.48 no.6
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• pp.348-353
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• 2015

In the present study, we investigated the characteristics of drug efflux transporter expressions following status epilepticus (SE). In the hippocampus and piriform cortex (PC), vasogenic edema peaked 3-4 days after SE. The expression of breast cancer resistance protein (BCRP), multidrug resistance protein-4 (MRP4), and p-glycoprotein (p-GP) were decreased 4 days after SE when vasogenic edema was peaked, but subsequently increased 4 weeks after SE. Multidrug resistance protein-1 (MRP1) expression gradually decreased in endothelial cells until 4 weeks after SE. These findings indicate that SE-induced vasogenic edema formation transiently reduced drug efflux pump expressions in endothelial cells. Subsequently, during recovery of vasogenic edema drug efflux pump expressions were differentially upregulated in astrocytes, neuropils, and endothelial cells. Therefore, we suggest that vasogenic edema formation may be a risk factor in pharmacoresistent epilepsy. [BMB Reports 2015; 48(6): 348-353]

• Korean Journal of Microbiology
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• v.49 no.2
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• pp.126-130
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• 2013

The major aim of this study was to investigate the distribution of genes that encode multidrug efflux pumps in Enterococci spp. isolates from bovine milk samples and antibiotic resistance patterns of these strains. Of the 245 isolates, 44.1% showed ampicillin resistance, 79.2% showed erythromycin resistance, 76.3% showed tetracycline resistance and 36.3% showed chloramphenicol resistance. In case of vancomycin and ciprofloxacin, all of the isolates were susceptible to these antibiotics. Of the 245 enterococcal isolates, 82.1% have MFS type eme(A) gene, 72.7% have ABC type efr(A) gene, 77.1% have ABC type efr(B) gene, and 71.8% have ABC type lsa gene. In case of Enterococcus faecalis, the original strain for these genes, 92.5% have eme(A), 87.4% have efr(A), 88.4% have efr(B), and 88.4% have lsa. Interestingly, in case of different species of Enterococci, eme(A) was also detected in four strains of E. faecium, seven strains of E. avium, four strains of E. durans and two strains of E. raffinosus. efr(A) was also detected in two strains of E. faecium and two strains of E. durans and efr(B) was also detected in four strains of E. faecium, five strains of E. avium and four strains of E. durans. This means the possibility of co-transfer of resistance genes between Enterococci species in natural environment. These results are the first report describing the presence of same multidrug efflux pumps in different species of Enterococci in Korea.