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Journal of Proteomics Sep 2022Carbapenem-resistant Klebsiella pneumoniae (CRKP) is an urgent threat to human health. Major outer membrane proteins (OMPs) porin mutation is one important resistance...
Carbapenem-resistant Klebsiella pneumoniae (CRKP) is an urgent threat to human health. Major outer membrane proteins (OMPs) porin mutation is one important resistance mechanism of CRKP, and may also affect the inhibition activity of β-lactam and β-lactamase inhibitor combinations. The ertapenem-resistant K. pneumoniae strain 2018B120 with major porin mutations was isolated from a clinical patient. Genomic and time-series proteomic analyses were conducted to retrieve the ertapenem-challenged response of 2018B120. The abundance changing of proteins from PTS systems, ABC transporters, the autoinducer 2 (AI-2) quorum sensing system, and antioxidant systems can be observed. Overexpression of alternative porins was also noticed to balance major porins' defection. These findings added a detailed regulation network in bacterial resistance mechanisms and gave new insights into bypass adaptation mechanisms the porin deficient bacteria adopted under carbapenem antibiotics pressure. SIGNIFICANCE: Outer membrane porins deficiency is an important mechanism of carbapenem resistance in K. pneumoniae. Comprehensive genomic and proteomic profiling of an ertapenem-resistant K. pneumoniae strain 2018B120 gives a detailed systematic regulation network in bacterial resistance mechanisms. Overexpression of alternative porins to balance major porins' defection was noticed, giving new insights into bypass adaptation mechanisms of porin deficient bacteria.
Topics: ATP-Binding Cassette Transporters; Anti-Bacterial Agents; Antioxidants; Bacterial Proteins; Carbapenems; Ertapenem; Humans; Klebsiella pneumoniae; Microbial Sensitivity Tests; Porins; Proteomics; beta-Lactam Resistance; beta-Lactamase Inhibitors; beta-Lactamases; beta-Lactams
PubMed: 36058541
DOI: 10.1016/j.jprot.2022.104715 -
Marine Drugs Apr 2018A diverse range of predatory marine gastropods produce toxins, yet most of these molecules remain uncharacterized. species have received the most attention from... (Review)
Review
A diverse range of predatory marine gastropods produce toxins, yet most of these molecules remain uncharacterized. species have received the most attention from researchers, leading to several conopeptides reaching clinical trials. This review aims to summarize what is known about bioactive compounds isolated from species of neglected marine gastropods, especially in the Turridae, Terebridae, Babyloniidae, Muricidae, Buccinidae, Colubrariidae, Nassariidae, Cassidae, and Ranellidae families. Multiple species have been reported to contain bioactive compounds with potential toxic activity, but most of these compounds have not been characterized or even clearly identified. The bioactive properties and potential applications of echotoxins and related porins from the Ranellidae family are discussed in more detail. Finally, the review concludes with a call for research on understudied species.
Topics: Animals; Aquatic Organisms; Biological Products; Biomedical Research; Biotechnology; Classification; Conotoxins; Conus Snail; Molecular Conformation; Porins; Predatory Behavior
PubMed: 29621159
DOI: 10.3390/md16040118 -
Current Protein & Peptide Science 2017Site-directed mutagenesis allows elucidation of the basic principles of the porin-driven membrane permeability and opens the possibility for the modulation of functional... (Review)
Review
Site-directed mutagenesis allows elucidation of the basic principles of the porin-driven membrane permeability and opens the possibility for the modulation of functional states of porin channels. The review is aimed to show the advantages of using mutant and chemically modified porins for obtaining detailed information about molecular mechanisms that underlie the non-specific transmembrane diffusion. We summarized data regarding the effects of the point substitutions and the external loop deletions on electrophysiological properties of general porins. The influence of charges inside the pore eyelet and the roles of external loops in ion conductance, ion selectivity, and voltage gating were described.
Topics: Bacterial Outer Membrane Proteins; Cell Membrane Permeability; Diffusion; Electrophysiological Phenomena; Mutagenesis, Site-Directed; Mutation; Porins
PubMed: 27593088
DOI: 10.2174/1389203717666160905145514 -
Nature Chemical Biology Dec 2020We live in the era of antibiotic resistance, and this problem will progressively worsen if no new solutions emerge. In particular, Gram-negative pathogens present both... (Review)
Review
We live in the era of antibiotic resistance, and this problem will progressively worsen if no new solutions emerge. In particular, Gram-negative pathogens present both biological and chemical challenges that hinder the discovery of new antibacterial drugs. First, these bacteria are protected from a variety of structurally diverse drugs by a low-permeability barrier composed of two membranes with distinct permeability properties, in addition to active drug efflux, making this cell envelope impermeable to most compounds. Second, chemical libraries currently used in drug discovery contain few compounds that can penetrate Gram-negative bacteria. As a result of these challenges, intensive screening campaigns have led to few successes, highlighting the need for new approaches to identify regions of chemical space that are specifically relevant to antibacterial drug discovery. Herein we provide an overview of emerging insights into this problem and outline a general approach to addressing it using prospective analysis of chemical libraries for the ability of compounds to accumulate in Gram-negative bacteria. The overall goal is to develop robust cheminformatic tools to predict Gram-negative permeation and efflux, which can then be used to guide medicinal chemistry campaigns and the design of antibacterial discovery libraries.
Topics: Anti-Bacterial Agents; Biological Transport; Cell Membrane; Cell Membrane Permeability; Cheminformatics; Chemistry, Pharmaceutical; Computer Simulation; Drug Discovery; Drug Resistance, Multiple, Bacterial; Gram-Negative Bacteria; Humans; Models, Statistical; Porins; Small Molecule Libraries; Structure-Activity Relationship
PubMed: 33199906
DOI: 10.1038/s41589-020-00674-6 -
Microbial Biotechnology May 2024Pseudomonas aeruginosa is a notorious multidrug-resistant pathogen that poses a serious and growing threat to the worldwide public health. The expression of resistance... (Review)
Review
Pseudomonas aeruginosa is a notorious multidrug-resistant pathogen that poses a serious and growing threat to the worldwide public health. The expression of resistance determinants is exquisitely modulated by the abundant regulatory proteins and the intricate signal sensing and transduction systems in this pathogen. Downregulation of antibiotic influx porin proteins and upregulation of antibiotic efflux pump systems owing to mutational changes in their regulators or the presence of distinct inducing molecular signals represent two of the most efficient mechanisms that restrict intracellular antibiotic accumulation and enable P. aeruginosa to resist multiple antibiotics. Treatment of P. aeruginosa infections is extremely challenging due to the highly inducible mechanism of antibiotic resistance. This review comprehensively summarizes the regulatory networks of the major porin proteins (OprD and OprH) and efflux pumps (MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY) that play critical roles in antibiotic influx and efflux in P. aeruginosa. It also discusses promising therapeutic approaches using safe and efficient adjuvants to enhance the efficacy of conventional antibiotics to combat multidrug-resistant P. aeruginosa by controlling the expression levels of porins and efflux pumps. This review not only highlights the complexity of the regulatory network that induces antibiotic resistance in P. aeruginosa but also provides important therapeutic implications in targeting the inducible mechanism of resistance.
Topics: Pseudomonas aeruginosa; Anti-Bacterial Agents; Gene Expression Regulation, Bacterial; Humans; Membrane Transport Proteins; Pseudomonas Infections; Drug Resistance, Multiple, Bacterial; Porins; Bacterial Proteins; Biological Transport
PubMed: 38801351
DOI: 10.1111/1751-7915.14487 -
Sub-cellular Biochemistry 2018Viroporins are short polypeptides encoded by viruses. These small membrane proteins assemble into oligomers that can permeabilize cellular lipid bilayers, disrupting the... (Review)
Review
Viroporins are short polypeptides encoded by viruses. These small membrane proteins assemble into oligomers that can permeabilize cellular lipid bilayers, disrupting the physiology of the host to the advantage of the virus. Consequently, efforts during the last few decades have been focused towards the discovery of viroporin channel inhibitors, but in general these have not been successful to produce licensed drugs. Viroporins are also involved in viral pathogenesis by engaging in critical interactions with viral proteins, or disrupting normal host cellular pathways through coordinated interactions with host proteins. These protein-protein interactions (PPIs) may become alternative attractive drug targets for the development of antivirals. In this sense, while thus far most antiviral molecules have targeted viral proteins, focus is moving towards targeting host proteins that are essential for virus replication. In principle, this largely would overcome the problem of resistance, with the possibility of using repositioned existing drugs. The precise role of these PPIs, their strain- and host- specificities, and the structural determination of the complexes involved, are areas that will keep the fields of virology and structural biology occupied for years to come. In the present review, we provide an update of the efforts in the characterization of the main PPIs for most viroporins, as well as the role of viroporins in these PPIs interactions.
Topics: Animals; Host-Pathogen Interactions; Humans; Porins; Viral Regulatory and Accessory Proteins; Virus Physiological Phenomena; Viruses
PubMed: 29900504
DOI: 10.1007/978-981-10-8456-0_15 -
BioEssays : News and Reviews in... Dec 2022Outer membrane proteins (OMPs) maintain the viability of Gram-negative bacteria by functioning as receptors, transporters, ion channels, lipases, and porins. Folding and...
Outer membrane proteins (OMPs) maintain the viability of Gram-negative bacteria by functioning as receptors, transporters, ion channels, lipases, and porins. Folding and assembly of OMPs involves synchronized action of chaperones and multi-protein machineries which escort the highly hydrophobic polypeptides to their target outer membrane in a folding competent state. Previous studies have identified proteins and their involvement along the OMP biogenesis pathway. Yet, the mechanisms of action and the intriguing ability of all these molecular machines to work without the typical cellular energy source of ATP, but solely based on thermodynamic principles, are still not well understood. Here, we highlight how different single-molecule studies can shed additional light on the mechanisms and kinetics of OMP biogenesis.
Topics: Bacterial Outer Membrane Proteins; Molecular Chaperones; Porins; Protein Folding; Gram-Negative Bacteria
PubMed: 36284497
DOI: 10.1002/bies.202200149 -
The Lancet. Microbe Dec 2021Bartonella bacilliformis is the aetiological agent of Carrión's disease, a biphasic and highly lethal illness formerly restricted to the South American Andes that is...
BACKGROUND
Bartonella bacilliformis is the aetiological agent of Carrión's disease, a biphasic and highly lethal illness formerly restricted to the South American Andes that is now spreading to adjacent areas. Reliable serodiagnostic approaches and vaccines are urgently needed. In this study, we aimed to identify immunodominant proteins of B bacilliformis and to establish novel and reliable serodiagnostic tools.
METHODS
We used a reverse vaccinology approach in combination with an analysis of heterologous genomic expression libraries to identify immunodominant proteins, on the basis of the genome sequences of B bacilliformis strains KC583 and KC584. Antigens were screened with serum samples collected from Peruvian patients with B bacilliformis infections and from German healthy blood donors without history of travel to South America. We further analysed immunoreactive proteins of B bacilliformis with immunoblotting and line blots. We used selected target proteins to develop a diagnostic ELISA. To assess the performance of this ELISA, we did receiver operating characteristic analyses to assess the area under the curve, cutoff values, sensitivities, and specificities with 95% CIs.
FINDINGS
We used serum samples obtained between Dec 23, 1990, and May 5, 2018, from 26 Peruvian patients with B bacilliformis infections and serum samples taken between Aug 28 and Aug 31, 2020, from 96 healthy German blood donors. 21 potentially immunodominant proteins were identified and recombinantly expressed, and their reactivity was assessed with immunoblotting and line blots. Of these 21 antigens, 14 were found to be immunoreactive. By using serum samples of Peruvian patients with Carrión's disease and of healthy German blood donors, we identified three antigens (porin B, autotransporter E, and hypothetical protein B) as suitable immunodominant antigens, and we applied them in a diagnostic ELISA using two different antigen combinations (porin B plus autotransporter E and porin B plus autotransporter E plus hypothetical protein B). For the combination of porin B and autotransporter E, with optical density measured at 450 nm (OD) cutoff value of 0·29, sensitivity was 80·8% (95% CI 60·7-93·5) and specificity was 94·8% (88·3-98·3) for all Peruvian patient samples. For a combination of porin B, autotransporter E, and hypothetical protein B, with an OD cutoff of 0·34, sensitivity was 76·9% (56·4-91·0) and specificity was 93·8% (86·9-97·7) for all Peruvian patient samples.
INTERPRETATION
This novel ELISA could represent a useful serodiagnostic tool for future epidemiological studies of B bacilliformis in endemic areas. Additionally, the immunodominant antigens we have identified could provide a first basis for future vaccine development to prevent the highly lethal Carrión's disease.
FUNDING
DRUID (Novel Drug Targets against Poverty-Related and Neglected Tropical Infectious Diseases) Initiative and Robert Koch Institute.
TRANSLATIONS
For the Spanish and Quechua translations of the abstract see Supplementary Materials section.
Topics: Bartonella Infections; Bartonella bacilliformis; Humans; Immunodominant Epitopes; Porins; Type V Secretion Systems
PubMed: 35544109
DOI: 10.1016/S2666-5247(21)00184-1 -
Methods (San Diego, Calif.) Aug 2016Single-molecule picometer resolution nanopore tweezers (SPRNT) is a new tool for analyzing the motion of nucleic acids through molecular motors. With SPRNT, individual... (Review)
Review
Single-molecule picometer resolution nanopore tweezers (SPRNT) is a new tool for analyzing the motion of nucleic acids through molecular motors. With SPRNT, individual enzymatic motions along DNA as small as 40pm can be resolved on sub-millisecond time scales. Additionally, SPRNT reveals an enzyme's exact location with respect to a DNA strand's nucleotide sequence, enabling identification of sequence-specific behaviors. SPRNT is enabled by a mutant version of the biological nanopore formed by Mycobacterium smegmatis porin A (MspA). SPRNT is strongly rooted in nanopore sequencing and therefore requires a solid understanding of basic principles of nanopore sequencing. Furthermore, SPRNT shares tools developed for nanopore sequencing and extends them to analysis of single-molecule kinetics. As such, this review begins with a brief history of our work developing the nanopore MspA for nanopore sequencing. We then describe the underlying principles of SPRNT, how it works in detail, and propose some potential future uses. We close with a comparison of SPRNT to other techniques and we present the methods that will enable others to use SPRNT.
Topics: Kinetics; Mycobacterium smegmatis; Nanopores; Nucleic Acids; Optical Tweezers; Porins; Single Molecule Imaging
PubMed: 27045943
DOI: 10.1016/j.ymeth.2016.03.026 -
Molecules (Basel, Switzerland) Jun 2021Here, we investigated general porin regulation in 488, the causative agent of Far Eastern scarlet-like fever, in response to sublethal concentrations of antibiotics. We...
Here, we investigated general porin regulation in 488, the causative agent of Far Eastern scarlet-like fever, in response to sublethal concentrations of antibiotics. We chose four antibiotics of different classes and measured gene expression using qRT-PCR and GFP reporter systems. Our data showed temporal regulation of the general porin genes and caused by antibiotic stress. The porin transcription initially decreased, providing early defensive response of the bacterium, while it returned to that of the untreated cells on prolonged antibiotic exposure. Unlike the major porin genes, the transcription of the alternative porin genes and was increased. Moreover, a short-term - and -mediated porin regulation was observed. The main finding was a phenotypic heterogeneity of population manifested in variable porin gene expression under carbenicillin exposure. This may offer adaptive fitness advantages for a particular bacterial subpopulation.
Topics: Anti-Bacterial Agents; Bacterial Proteins; Carbenicillin; Gene Expression Regulation, Bacterial; Porins; Stress, Physiological; Yersinia pseudotuberculosis
PubMed: 34203552
DOI: 10.3390/molecules26133956