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Monitoring antibacterial permeabilization in real time using time-resolved flow cytometry

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dc.contributor.authorFreire, João Miguel
dc.contributor.authorGaspar, Diana
dc.contributor.authorTorre, Beatriz Garcia de la
dc.contributor.authorVeiga, Ana Salomé
dc.contributor.authorAndreu, David
dc.contributor.authorCastanho, Miguel A. R. B.
dc.date.accessioned2016-04-18T16:01:06Z
dc.date.available2016-04-18T16:01:06Z
dc.date.issued2015
dc.description© 2014 Elsevier B.V. All rights reservedpt_PT
dc.description.abstractDespite the intensive study of antibiotic-induced bacterial permeabilization, its kinetics and molecular mechanism remain largely elusive. A new methodology that extends the concept of the live–dead assay in flow cytometry to real time-resolved detection was used to overcome these limitations. The antimicrobial activity of pepR was monitored in time-resolved flow cytometry for three bacterial strains: Escherichia coli (ATCC 25922), E. coli K-12 (CGSC Strain 4401) and E. coli JW3596-1 (CGSC Strain 11805). The latter strain has truncated lipopolysaccharides (LPS) in the outer membrane. This new methodology provided information on the efficacy of the antibiotics and sheds light on their mode of action atmembrane-level. Kinetic data regarding antibiotic binding and lytic actionwere retrieved.Membrane interaction and permeabilization events differ significantly among strains. The truncation of LPS moieties does not hamper AMP binding but compromises membrane disruption and bacterial killing. We demonstrated the usefulness of time-resolved flow cytometry to study antimicrobial-induced permeabilization by collecting kinetic data that contribute to characterize the action of antibiotics directly on bacteria.pt_PT
dc.description.sponsorshipWe thank CGSC, Coli Genetic Stock Center, for the K-12 and JW3596-1 Escherichia coli strains. This work was supported by project PTDC/QUIBIQ/112929/2009 from Fundação para a Ciência e a Tecnologia–Ministério da Educação e Ciência (FCT-MEC, Portugal). JMF and DG also acknowledge FCT-MEC for fellowships SFRH/BD/70423/2010 and SFRH/BPD/73500/2010, respectively, and ASV for funding within the FCT Investigator Programme (IF/00803/2012). Work at Pompeu Fabra University was supported by grants from the Spanish Ministry of Economy and Competitiveness (SAF2011-24899) and from Generalitat de Catalunya (SGR2009-0492).pt_PT
dc.identifier.citationBiochimica et Biophysica Acta 1848 (2015) 554–560pt_PT
dc.identifier.doi10.1016/j.bbamem.2014.11.001pt_PT
dc.identifier.issn0005-2736
dc.identifier.urihttp://hdl.handle.net/10451/23430
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherElsevierpt_PT
dc.relationSTUDYING AND DEVELOPING ANTIMICROBIAL PEPTIDE HYDROGELS USED IN TISSUE REGENERATION
dc.relation.publisherversionhttp://www.sciencedirect.com/science/article/pii/S0005273614003770pt_PT
dc.subjectTime-resolved flow cytometrypt_PT
dc.subjectAntimicrobial peptidept_PT
dc.subjectLive/dead assaypt_PT
dc.subjectPeptide–lipid interactionpt_PT
dc.subjectAntimicrobial actionpt_PT
dc.subjectLipopolysaccharidept_PT
dc.titleMonitoring antibacterial permeabilization in real time using time-resolved flow cytometrypt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.awardTitleSTUDYING AND DEVELOPING ANTIMICROBIAL PEPTIDE HYDROGELS USED IN TISSUE REGENERATION
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FQUI-BIQ%2F112929%2F2009/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/SFRH/SFRH%2FBD%2F70423%2F2010/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT//SFRH%2FBPD%2F73500%2F2010/PT
oaire.citation.titleBiochimica et Biophysica Acta (BBA) - Biomembranespt_PT
oaire.fundingStream3599-PPCDT
oaire.fundingStreamSFRH
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
rcaap.rightsclosedAccesspt_PT
rcaap.typearticlept_PT
relation.isProjectOfPublication954cb0bb-03f8-45f6-a48d-27c88a2f8690
relation.isProjectOfPublication1a004e63-0ace-4b41-bd0c-db90b919807e
relation.isProjectOfPublication8aef3bd6-260c-4cad-afd0-15952a73f08e
relation.isProjectOfPublication.latestForDiscovery1a004e63-0ace-4b41-bd0c-db90b919807e

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