Real-Time Probing of Membrane Transport in Living Microbial Cells Using Single Nanoparticle Optics and Living Cell Imaging
Membrane transport plays a leading role in a wide spectrum of cellular and subcellular pathways, including multidrug resistance (MDR), cellular signaling, and cell−cell communication. Pseudomonas aeruginosa is renowned for its intriguing membrane transport mechanisms, such as the interplay of membra...
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Veröffentlicht in: | Biochemistry (Easton) 2004-08, Vol.43 (32), p.10400-10413 |
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description | Membrane transport plays a leading role in a wide spectrum of cellular and subcellular pathways, including multidrug resistance (MDR), cellular signaling, and cell−cell communication. Pseudomonas aeruginosa is renowned for its intriguing membrane transport mechanisms, such as the interplay of membrane permeability and extrusion machinery, leading to selective accumulation of specific intracellular substances and MDR. Despite extensive studies, the mechanisms of membrane transport in living microbial cells remain incompletely understood. In this study, we directly measure real-time change of membrane permeability and pore sizes of P. aeruginosa at the nanometer scale using the intrinsic color index (surface plasmon resonance spectra) of silver (Ag) nanoparticles as the nanometer size index probes. The results show that Ag nanoparticles with sizes ranging up to 80 nm are accumulated in living microbial cells, demonstrating that these Ag nanoparticles transport through the inner and outer membrane of the cells. In addition, a greater number of larger intracellular Ag nanoparticles are observed in the cells as chloramphenicol concentration increases, suggesting that chloramphenicol increases membrane permeability and porosity. Furthermore, studies of mutants (nalB-1 and ΔABM) show that the accumulation rate of intracellular Ag nanoparticles depends on the expression level of the extrusion pump (MexAB−OprM), suggesting that the extrusion pump plays an important role in controlling the accumulation of Ag nanoparticles in living cells. Moreover, the accumulation kinetics measured by Ag nanoparticles are similar to those measured using a small fluorescent molecule (EtBr), eliminating the possibility of steric and size effects of Ag nanoparticle probes. Susceptibility measurements also suggest that a low concentration of Ag nanoparticles (1.3 pM) does not create significant toxicity for the cells, further validating that single Ag nanoparticles (1.3 pM) can be used as biocompatible nanoprobes for the study of membrane transport kinetics in living microbial cells. |
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Pseudomonas aeruginosa is renowned for its intriguing membrane transport mechanisms, such as the interplay of membrane permeability and extrusion machinery, leading to selective accumulation of specific intracellular substances and MDR. Despite extensive studies, the mechanisms of membrane transport in living microbial cells remain incompletely understood. In this study, we directly measure real-time change of membrane permeability and pore sizes of P. aeruginosa at the nanometer scale using the intrinsic color index (surface plasmon resonance spectra) of silver (Ag) nanoparticles as the nanometer size index probes. The results show that Ag nanoparticles with sizes ranging up to 80 nm are accumulated in living microbial cells, demonstrating that these Ag nanoparticles transport through the inner and outer membrane of the cells. In addition, a greater number of larger intracellular Ag nanoparticles are observed in the cells as chloramphenicol concentration increases, suggesting that chloramphenicol increases membrane permeability and porosity. Furthermore, studies of mutants (nalB-1 and ΔABM) show that the accumulation rate of intracellular Ag nanoparticles depends on the expression level of the extrusion pump (MexAB−OprM), suggesting that the extrusion pump plays an important role in controlling the accumulation of Ag nanoparticles in living cells. Moreover, the accumulation kinetics measured by Ag nanoparticles are similar to those measured using a small fluorescent molecule (EtBr), eliminating the possibility of steric and size effects of Ag nanoparticle probes. Susceptibility measurements also suggest that a low concentration of Ag nanoparticles (1.3 pM) does not create significant toxicity for the cells, further validating that single Ag nanoparticles (1.3 pM) can be used as biocompatible nanoprobes for the study of membrane transport kinetics in living microbial cells.</description><identifier>ISSN: 0006-2960</identifier><identifier>EISSN: 1520-4995</identifier><identifier>DOI: 10.1021/bi036231a</identifier><identifier>PMID: 15301539</identifier><language>eng</language><publisher>United States: American Chemical Society</publisher><subject>Anti-Bacterial Agents - pharmacology ; Bacterial Outer Membrane Proteins - metabolism ; Bacterial Proteins - genetics ; Bacterial Proteins - metabolism ; Biological Transport ; Cell Membrane - metabolism ; Cell Membrane Permeability ; Chloramphenicol - pharmacology ; Drug Resistance, Bacterial ; Ethidium - chemistry ; Kinetics ; Membrane Transport Proteins - metabolism ; Microbial Sensitivity Tests ; Optics and Photonics ; Particle Size ; Pseudomonas aeruginosa ; Pseudomonas aeruginosa - cytology ; Pseudomonas aeruginosa - drug effects ; Pseudomonas aeruginosa - metabolism ; Silver Staining ; Surface Plasmon Resonance</subject><ispartof>Biochemistry (Easton), 2004-08, Vol.43 (32), p.10400-10413</ispartof><rights>Copyright © 2004 American Chemical Society</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a446t-d083d53b7b82703ff1b2c5c0abaa8407e8cd0f9bf445488ee9a3cb28154af6df3</citedby><cites>FETCH-LOGICAL-a446t-d083d53b7b82703ff1b2c5c0abaa8407e8cd0f9bf445488ee9a3cb28154af6df3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/bi036231a$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/bi036231a$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,776,780,2752,27053,27901,27902,56713,56763</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15301539$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Xu, Xiao-Hong Nancy</creatorcontrib><creatorcontrib>Brownlow, William J</creatorcontrib><creatorcontrib>Kyriacou, Sophia V</creatorcontrib><creatorcontrib>Wan, Qian</creatorcontrib><creatorcontrib>Viola, Joshua J</creatorcontrib><title>Real-Time Probing of Membrane Transport in Living Microbial Cells Using Single Nanoparticle Optics and Living Cell Imaging</title><title>Biochemistry (Easton)</title><addtitle>Biochemistry</addtitle><description>Membrane transport plays a leading role in a wide spectrum of cellular and subcellular pathways, including multidrug resistance (MDR), cellular signaling, and cell−cell communication. Pseudomonas aeruginosa is renowned for its intriguing membrane transport mechanisms, such as the interplay of membrane permeability and extrusion machinery, leading to selective accumulation of specific intracellular substances and MDR. Despite extensive studies, the mechanisms of membrane transport in living microbial cells remain incompletely understood. In this study, we directly measure real-time change of membrane permeability and pore sizes of P. aeruginosa at the nanometer scale using the intrinsic color index (surface plasmon resonance spectra) of silver (Ag) nanoparticles as the nanometer size index probes. The results show that Ag nanoparticles with sizes ranging up to 80 nm are accumulated in living microbial cells, demonstrating that these Ag nanoparticles transport through the inner and outer membrane of the cells. In addition, a greater number of larger intracellular Ag nanoparticles are observed in the cells as chloramphenicol concentration increases, suggesting that chloramphenicol increases membrane permeability and porosity. Furthermore, studies of mutants (nalB-1 and ΔABM) show that the accumulation rate of intracellular Ag nanoparticles depends on the expression level of the extrusion pump (MexAB−OprM), suggesting that the extrusion pump plays an important role in controlling the accumulation of Ag nanoparticles in living cells. Moreover, the accumulation kinetics measured by Ag nanoparticles are similar to those measured using a small fluorescent molecule (EtBr), eliminating the possibility of steric and size effects of Ag nanoparticle probes. Susceptibility measurements also suggest that a low concentration of Ag nanoparticles (1.3 pM) does not create significant toxicity for the cells, further validating that single Ag nanoparticles (1.3 pM) can be used as biocompatible nanoprobes for the study of membrane transport kinetics in living microbial cells.</description><subject>Anti-Bacterial Agents - pharmacology</subject><subject>Bacterial Outer Membrane Proteins - metabolism</subject><subject>Bacterial Proteins - genetics</subject><subject>Bacterial Proteins - metabolism</subject><subject>Biological Transport</subject><subject>Cell Membrane - metabolism</subject><subject>Cell Membrane Permeability</subject><subject>Chloramphenicol - pharmacology</subject><subject>Drug Resistance, Bacterial</subject><subject>Ethidium - chemistry</subject><subject>Kinetics</subject><subject>Membrane Transport Proteins - metabolism</subject><subject>Microbial Sensitivity Tests</subject><subject>Optics and Photonics</subject><subject>Particle Size</subject><subject>Pseudomonas aeruginosa</subject><subject>Pseudomonas aeruginosa - cytology</subject><subject>Pseudomonas aeruginosa - drug effects</subject><subject>Pseudomonas aeruginosa - metabolism</subject><subject>Silver Staining</subject><subject>Surface Plasmon Resonance</subject><issn>0006-2960</issn><issn>1520-4995</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNptkE1PGzEQhq2KqgTaA38A-QISh23ttb3rPULUpJFCQSVRerPGuzYy7Bf2pmr59XiVFC4cPKPX88zY8yJ0QslXSlL6TTvCspRR-IAmVKQk4UUhDtCEEJIlaZGRQ3QUwkOUnOT8EzqkgpF4igl6_mWgTlauMfjWd9q197iz-No02kNr8CrG0Hd-wK7FS_dnrF-7ciShxlNT1wGvw3h7F0Nt8E9oux784MoobvqYA4a2-t87duBFA_dRfEYfLdTBfNnnY7SefV9NfyTLm_lierlMgPNsSCoiWSWYzrVMc8KspTotRUlAA8i4j5FlRWyhLeeCS2lMAazUqaSCg80qy47R-W5u77unrQmDalwo40figt02KCqJoJLlEbzYgXG_ELyxqveuAf9PUaJGo9Wr0ZE93Q_d6sZUb-Te2QgkO8CFwfx9rYN_VFnOcqFWt3dqVuSb-W-xUVeRP9vxUAb10G19Gz155-EXfNGUbA</recordid><startdate>20040817</startdate><enddate>20040817</enddate><creator>Xu, Xiao-Hong Nancy</creator><creator>Brownlow, William J</creator><creator>Kyriacou, Sophia V</creator><creator>Wan, Qian</creator><creator>Viola, Joshua J</creator><general>American Chemical Society</general><scope>BSCLL</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7T7</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>P64</scope></search><sort><creationdate>20040817</creationdate><title>Real-Time Probing of Membrane Transport in Living Microbial Cells Using Single Nanoparticle Optics and Living Cell Imaging</title><author>Xu, Xiao-Hong Nancy ; Brownlow, William J ; Kyriacou, Sophia V ; Wan, Qian ; Viola, Joshua J</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a446t-d083d53b7b82703ff1b2c5c0abaa8407e8cd0f9bf445488ee9a3cb28154af6df3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2004</creationdate><topic>Anti-Bacterial Agents - pharmacology</topic><topic>Bacterial Outer Membrane Proteins - metabolism</topic><topic>Bacterial Proteins - genetics</topic><topic>Bacterial Proteins - metabolism</topic><topic>Biological Transport</topic><topic>Cell Membrane - metabolism</topic><topic>Cell Membrane Permeability</topic><topic>Chloramphenicol - pharmacology</topic><topic>Drug Resistance, Bacterial</topic><topic>Ethidium - chemistry</topic><topic>Kinetics</topic><topic>Membrane Transport Proteins - metabolism</topic><topic>Microbial Sensitivity Tests</topic><topic>Optics and Photonics</topic><topic>Particle Size</topic><topic>Pseudomonas aeruginosa</topic><topic>Pseudomonas aeruginosa - cytology</topic><topic>Pseudomonas aeruginosa - drug effects</topic><topic>Pseudomonas aeruginosa - metabolism</topic><topic>Silver Staining</topic><topic>Surface Plasmon Resonance</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Xu, Xiao-Hong Nancy</creatorcontrib><creatorcontrib>Brownlow, William J</creatorcontrib><creatorcontrib>Kyriacou, Sophia V</creatorcontrib><creatorcontrib>Wan, Qian</creatorcontrib><creatorcontrib>Viola, Joshua J</creatorcontrib><collection>Istex</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><jtitle>Biochemistry (Easton)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Xu, Xiao-Hong Nancy</au><au>Brownlow, William J</au><au>Kyriacou, Sophia V</au><au>Wan, Qian</au><au>Viola, Joshua J</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Real-Time Probing of Membrane Transport in Living Microbial Cells Using Single Nanoparticle Optics and Living Cell Imaging</atitle><jtitle>Biochemistry (Easton)</jtitle><addtitle>Biochemistry</addtitle><date>2004-08-17</date><risdate>2004</risdate><volume>43</volume><issue>32</issue><spage>10400</spage><epage>10413</epage><pages>10400-10413</pages><issn>0006-2960</issn><eissn>1520-4995</eissn><abstract>Membrane transport plays a leading role in a wide spectrum of cellular and subcellular pathways, including multidrug resistance (MDR), cellular signaling, and cell−cell communication. Pseudomonas aeruginosa is renowned for its intriguing membrane transport mechanisms, such as the interplay of membrane permeability and extrusion machinery, leading to selective accumulation of specific intracellular substances and MDR. Despite extensive studies, the mechanisms of membrane transport in living microbial cells remain incompletely understood. In this study, we directly measure real-time change of membrane permeability and pore sizes of P. aeruginosa at the nanometer scale using the intrinsic color index (surface plasmon resonance spectra) of silver (Ag) nanoparticles as the nanometer size index probes. The results show that Ag nanoparticles with sizes ranging up to 80 nm are accumulated in living microbial cells, demonstrating that these Ag nanoparticles transport through the inner and outer membrane of the cells. In addition, a greater number of larger intracellular Ag nanoparticles are observed in the cells as chloramphenicol concentration increases, suggesting that chloramphenicol increases membrane permeability and porosity. Furthermore, studies of mutants (nalB-1 and ΔABM) show that the accumulation rate of intracellular Ag nanoparticles depends on the expression level of the extrusion pump (MexAB−OprM), suggesting that the extrusion pump plays an important role in controlling the accumulation of Ag nanoparticles in living cells. Moreover, the accumulation kinetics measured by Ag nanoparticles are similar to those measured using a small fluorescent molecule (EtBr), eliminating the possibility of steric and size effects of Ag nanoparticle probes. Susceptibility measurements also suggest that a low concentration of Ag nanoparticles (1.3 pM) does not create significant toxicity for the cells, further validating that single Ag nanoparticles (1.3 pM) can be used as biocompatible nanoprobes for the study of membrane transport kinetics in living microbial cells.</abstract><cop>United States</cop><pub>American Chemical Society</pub><pmid>15301539</pmid><doi>10.1021/bi036231a</doi><tpages>14</tpages></addata></record> |
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subjects | Anti-Bacterial Agents - pharmacology Bacterial Outer Membrane Proteins - metabolism Bacterial Proteins - genetics Bacterial Proteins - metabolism Biological Transport Cell Membrane - metabolism Cell Membrane Permeability Chloramphenicol - pharmacology Drug Resistance, Bacterial Ethidium - chemistry Kinetics Membrane Transport Proteins - metabolism Microbial Sensitivity Tests Optics and Photonics Particle Size Pseudomonas aeruginosa Pseudomonas aeruginosa - cytology Pseudomonas aeruginosa - drug effects Pseudomonas aeruginosa - metabolism Silver Staining Surface Plasmon Resonance |
title | Real-Time Probing of Membrane Transport in Living Microbial Cells Using Single Nanoparticle Optics and Living Cell Imaging |
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