scholarly journals Role of Porins for Uptake of Antibiotics by Mycobacterium smegmatis

2008 ◽  
Vol 52 (9) ◽  
pp. 3127-3134 ◽  
Author(s):  
Olga Danilchanka ◽  
Mikhail Pavlenok ◽  
Michael Niederweis
Keyword(s):  
Outer Membrane ◽  
Mycobacterium Smegmatis ◽  
Lipid Membrane ◽  
Model Organism ◽  
Permeability Barrier ◽  
Opportunistic Pathogens ◽  
Transport Pathways ◽  
Antibiotics Use ◽  
Outer Membrane Permeability

ABSTRACT The outer membrane of mycobacteria presents an effective permeability barrier for many antibiotics. Transport pathways across this membrane are unknown for most drugs. Here, we examined which antibiotics utilize the porin pathway across the outer membrane of the model organism Mycobacterium smegmatis. Deletion of the porins MspA and MspC drastically increased the resistance of M. smegmatis ML10 to β-lactam antibiotics, while its β-lactamase activity remained unchanged. These results are consistent with the ninefold-reduced outer membrane permeability of the M. smegmatis porin mutants for cephaloridine and strongly indicate that β-lactam antibiotics rely on the porin pathway. The porin mutant ML10 accumulated less chloramphenicol and norfloxacin and was less susceptible to these antibiotics than wild-type M. smegmatis. These results demonstrated that small and hydrophilic antibiotics use the Msp porins for entering the cell. In contrast to norfloxacin, the hydrophobic moxifloxacin was 32-fold more effective in inhibiting the growth of M. smegmatis, presumably because it was able to diffuse through the lipid membrane. Structural models indicated that erythromycin, kanamycin, and vancomycin are too large to move through the MspA channel. This study presents the first experimental evidence that hydrophilic fluoroquinolones and chloramphenicol diffuse through porins in mycobacteria. Thus, mutations resulting in less efficient porins or lower porin expression levels are likely to represent a mechanism for the opportunistic pathogens M. avium, M. chelonae, and M. fortuitum, which have Msp-like porins, to acquire resistance to fluoroquinolones.

scholarly journals Multidrug Resistance of a Porin Deletion Mutant of Mycobacterium smegmatis

2004 ◽  
Vol 48 (11) ◽  
pp. 4163-4170 ◽  
Author(s):  
Joachim Stephan ◽  
Claudia Mailaender ◽  
Gilles Etienne ◽  
Mamadou Daffé ◽  
Michael Niederweis
Keyword(s):  
Multidrug Resistance ◽  
Outer Membrane ◽  
Mycobacterium Smegmatis ◽  
Antibiotic Sensitivity ◽  
Chemotherapeutic Agents ◽  
Intrinsic Resistance ◽  
Successful Design ◽  
High Level ◽  
Outer Membrane Permeability

ABSTRACT Mycobacteria contain an outer membrane of unusually low permeability which contributes to their intrinsic resistance to many agents. It is assumed that small and hydrophilic antibiotics cross the outer membrane via porins, whereas hydrophobic antibiotics may diffuse through the membrane directly. A mutant of Mycobacterium smegmatis lacking the major porin MspA was used to examine the role of the porin pathway in antibiotic sensitivity. Deletion of the mspA gene caused high-level resistance of M. smegmatis to 256 μg of ampicillin/ml by increasing the MIC 16-fold. The permeation of cephaloridine in the mspA mutant was reduced ninefold, and the resistance increased eightfold. This established a clear relationship between the activity and the outer membrane permeation of cephaloridine. Surprisingly, the MICs of the large and/or hydrophobic antibiotics vancomycin, erythromycin, and rifampin for the mspA mutant were increased 2- to 10-fold. This is in contrast to those for Escherichia coli, whose sensitivity to these agents was not affected by deletion of porin genes. Uptake of the very hydrophobic steroid chenodeoxycholate by the mspA mutant was retarded threefold, which supports the hypothesis that loss of MspA indirectly reduces the permeability by the lipid pathway. The multidrug resistance of the mspA mutant highlights the prominent role of outer membrane permeability for the sensitivity of M. smegmatis to antibiotics. An understanding of the pathways across the outer membrane is essential to the successful design of chemotherapeutic agents with activities against mycobacteria.

scholarly journals An “Uncommon” Role for Cyclic Enterobacterial Common Antigen in Maintaining Outer Membrane Integrity

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Corey S. Westfall
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Membrane Integrity ◽  
Physiological Role ◽  
Permeability Barrier ◽  
Common Antigen ◽  
Enterobacterial Common Antigen ◽  
Inner Membrane Protein ◽  
Outer Membrane Permeability

ABSTRACTAlthough discovered over 50 years ago, the physiological role of enterobacterial common antigen, a surface antigen produced by all members of theEnterobacteriaceae, has been poorly understood. In the work of Mitchell et al. (mBio 9:e01321-18, 2018,https://doi.org/10.1128/mBio.01321-18), the cyclized version of enterobacterial common antigen has been shown to play a role in maintaining the outer membrane permeability barrier, possibly through the inner membrane protein YhdP. This work also provides the tests needed to separate true effects from the numerous possible artifacts possible with mutations in enterobacterial common antigen synthesis.

scholarly journals Molecular Basis of Bacterial Outer Membrane Permeability Revisited

2003 ◽  
Vol 67 (4) ◽  
pp. 593-656 ◽  
Author(s):  
Hiroshi Nikaido
Keyword(s):  
Outer Membrane ◽  
Pathogenic Bacteria ◽  
Structural Knowledge ◽  
Permeability Barrier ◽  
Lipophilic Compounds ◽  
Bacterial Outer Membrane ◽  
Membrane Components ◽  
Outer Membrane Permeability ◽  
Protein Channels

SUMMARY Gram-negative bacteria characteristically are surrounded by an additional membrane layer, the outer membrane. Although outer membrane components often play important roles in the interaction of symbiotic or pathogenic bacteria with their host organisms, the major role of this membrane must usually be to serve as a permeability barrier to prevent the entry of noxious compounds and at the same time to allow the influx of nutrient molecules. This review summarizes the development in the field since our previous review (H. Nikaido and M. Vaara, Microbiol. Rev. 49:1-32, 1985) was published. With the discovery of protein channels, structural knowledge enables us to understand in molecular detail how porins, specific channels, TonB-linked receptors, and other proteins function. We are now beginning to see how the export of large proteins occurs across the outer membrane. With our knowledge of the lipopolysaccharide-phospholipid asymmetric bilayer of the outer membrane, we are finally beginning to understand how this bilayer can retard the entry of lipophilic compounds, owing to our increasing knowledge about the chemistry of lipopolysaccharide from diverse organisms and the way in which lipopolysaccharide structure is modified by environmental conditions.

scholarly journals Outer Membrane Permeability Barrier in Escherichia coli Mutants That Are Defective in the Late Acyltransferases of Lipid A Biosynthesis

1999 ◽  
Vol 43 (6) ◽  
pp. 1459-1462 ◽  
Author(s):  
Martti Vaara ◽  
Marjatta Nurminen
Keyword(s):  
Escherichia Coli ◽  
Membrane Permeability ◽  
Outer Membrane ◽  
Lipid A ◽  
Enteric Bacteria ◽  
Normal Function ◽  
Permeability Barrier ◽  
Core Oligosaccharide ◽  
Hydrophobic Solutes ◽  
Outer Membrane Permeability

ABSTRACT The tight packing of six fatty acids in the lipid A constituent of lipopolysaccharide (LPS) has been proposed to contribute to the unusually low permeability of the outer membrane of gram-negative enteric bacteria to hydrophobic antibiotics. Here it is shown that theEscherichia coli msbB mutant, which elaborates defective, penta-acylated lipid A, is practically as resistant to a representative set of hydrophobic solutes (rifampin, fusidic acid, erythromycin, clindamycin, and azithromycin) as the parent-type control strain. The susceptibility index, i.e., the approximate ratio between the MIC for the msbB mutant and that for the parent-type control, was maximally 2.7-fold. In comparison, the rfa mutant defective in the deep core oligosaccharide part of LPS displayed indices ranging from 20 to 64. The lpxA and lpxD lipid A mutants had indices higher than 512. Furthermore, the msbBmutant was resistant to glycopeptides (vancomycin, teicoplanin), whereas the rfa, lpxA, and lpxDmutants were susceptible. The msbB htrB double mutant, which elaborates even-more-defective, partially tetra-acylated lipid A, was still less susceptible than the rfa mutant. These findings indicate that hexa-acylated lipid A is not a prerequisite for the normal function of the outer membrane permeability barrier.

scholarly journals Essential Role of the ESX-5 Secretion System in Outer Membrane Permeability of Pathogenic Mycobacteria

PLoS Genetics ◽  
2015 ◽  
Vol 11 (5) ◽  
pp. e1005190 ◽  
Author(s):  
Louis S. Ates ◽  
Roy Ummels ◽  
Susanna Commandeur ◽  
Robert van der Weerd ◽  
Marion Sparrius ◽  
...  
Keyword(s):  
Membrane Permeability ◽  
Outer Membrane ◽  
Essential Role ◽  
Secretion System ◽  
Outer Membrane Permeability

scholarly journals Importance of Real-Time Assays To Distinguish Multidrug Efflux Pump-Inhibiting and Outer Membrane-Destabilizing Activities in Escherichia coli

2015 ◽  
Vol 197 (15) ◽  
pp. 2479-2488 ◽  
Author(s):  
Rajeev Misra ◽  
Keith D. Morrison ◽  
Hyun Jae Cho ◽  
Thanh Khuu
Keyword(s):  
Escherichia Coli ◽  
Membrane Permeability ◽  
Outer Membrane ◽  
Real Time ◽  
Efflux Pump ◽  
Membrane Integrity ◽  
Efflux Pumps ◽  
Permeability Barrier ◽  
Multidrug Efflux ◽  
Outer Membrane Permeability

ABSTRACTThe constitutively expressed AcrAB multidrug efflux system ofEscherichia colishows a high degree of homology with the normally silent AcrEF system. Exposure of a strain withacrABdeleted to antibiotic selection pressure frequently leads to the insertion sequence-mediated activation of the homologous AcrEF system. In this study, we used strains constitutively expressing either AcrAB or AcrEF from their normal chromosomal locations to resolve a controversy about whether phenylalanylarginine β-naphthylamide (PAβN) inhibits the activities of AcrAB and AcrEF and/or acts synergistically with antibiotics by destabilizing the outer membrane permeability barrier. Real-time efflux assays allowed a clear distinction between the efflux pump-inhibiting activity of PAβN and the outer membrane-destabilizing action of polymyxin B nonapeptide (PMXBN). When added in equal amounts, PAβN, but not PMXBN, strongly inhibited the efflux activities of both AcrAB and AcrEF pumps. In contrast, when outer membrane destabilization was assessed by the nitrocefin hydrolysis assay, PMXBN exerted a much greater damaging effect than PAβN. Strong action of PAβN in inhibiting efflux activity compared to its weak action in destabilizing the outer membrane permeability barrier suggests that PAβN acts mainly by inhibiting efflux pumps. We concluded that at low concentrations, PAβN acts specifically as an inhibitor of both AcrAB and AcrEF efflux pumps; however, at high concentrations, PAβN in the efflux-proficient background not only inhibits efflux pump activity but also destabilizes the membrane. The effects of PAβN on membrane integrity are compounded in cells unable to extrude PAβN.IMPORTANCEThe increase in multidrug-resistant bacterial pathogens at an alarming rate has accelerated the need for implementation of better antimicrobial stewardship, discovery of new antibiotics, and deeper understanding of the mechanism of drug resistance. The work carried out in this study highlights the importance of employing real-time fluorescence-based assays in differentiating multidrug efflux-inhibitory and outer membrane-destabilizing activities of antibacterial compounds.

scholarly journals Multivesicular Bodies as a Platform for Formation of the Marburg Virus Envelope

2004 ◽  
Vol 78 (22) ◽  
pp. 12277-12287 ◽  
Author(s):  
Larissa Kolesnikova ◽  
Beate Berghöfer ◽  
Sandra Bamberg ◽  
Stephan Becker
Keyword(s):  
Intracellular Transport ◽  
Matrix Protein ◽  
Lipid Membrane ◽  
Marburg Virus ◽  
Viral Envelope ◽  
Multivesicular Bodies ◽  
Virus Envelope ◽  
Transport Pathways ◽  
Virus Like Particles

ABSTRACT The Marburg virus (MARV) envelope consists of a lipid membrane and two major proteins, the matrix protein VP40 and the glycoprotein GP. Both proteins use different intracellular transport pathways: GP utilizes the exocytotic pathway, while VP40 is transported through the retrograde late endosomal pathway. It is currently unknown where the proteins combine to form the viral envelope. In the present study, we identified the intracellular site where the two major envelope proteins of MARV come together as peripheral multivesicular bodies (MVBs). Upon coexpression with VP40, GP is redistributed from the trans-Golgi network into the VP40-containing MVBs. Ultrastructural analysis of MVBs suggested that they provide the platform for the formation of membrane structures that bud as virus-like particles from the cell surface. The virus-like particles contain both VP40 and GP. Single expression of GP also resulted in the release of particles, which are round or pleomorphic. Single expression of VP40 led to the release of filamentous structures that closely resemble viral particles and contain traces of endosomal marker proteins. This finding indicated a central role of VP40 in the formation of the filamentous structure of MARV particles, which is similar to the role of the related Ebola virusVP40. In MARV-infected cells, VP40 and GP are colocalized in peripheral MVBs as well. Moreover, intracellular budding of progeny virions into MVBs was frequently detected. Taken together, these results demonstrate an intracellular intersection between GP and VP40 pathways and suggest a crucial role of the late endosomal compartment for the formation of the viral envelope.

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