Abstract
We investigated the transmembrane movement of fluorescent labeled phospholipids in inverted inner membrane vesicles (IIMV) of Escherichia coli (E. coli) wild-type strain (MG1655), as well as in proteoliposomes reconstituted from detergent extracts of the IIMV. The transbilayer movement of 1-myristoyl-2-[6-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]caproyl]-sn-glycero-3-phosphoethanolamine (M-C6-NBD-PE) and -phosphocholine (M-C6-NBD-PC) was measured by a fluorescence stopped-flow back-exchange assay. Both analogues were rapidly translocated across the IIMV membrane, with half-times of <1 min (outward movement) and approximately 3 min (inward movement). No flip-flop was detected in protein-free liposomes, but in IIMV-derived proteoliposomes flip-flop of M-C6-NBD-PE occurred similarly to IIMV and could be largely eliminated by proteinase K treatment.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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4-Chloro-7-nitrobenzofurazan / analogs & derivatives*
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4-Chloro-7-nitrobenzofurazan / chemistry
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4-Chloro-7-nitrobenzofurazan / metabolism
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Biological Transport
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Carrier Proteins / metabolism
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Cytoplasm / chemistry
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Cytoplasm / metabolism*
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Detergents
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Endopeptidase K / metabolism
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Escherichia coli / chemistry
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Escherichia coli / metabolism*
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Fluorescent Dyes / chemistry
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Fluorescent Dyes / metabolism*
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Intracellular Membranes / chemistry
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Intracellular Membranes / metabolism*
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Kinetics
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Lipid Bilayers / chemistry
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Lipid Bilayers / metabolism*
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Membrane Proteins / metabolism
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Phosphatidylcholines / chemistry
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Phosphatidylcholines / metabolism
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Phosphatidylethanolamines / chemistry
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Phosphatidylethanolamines / metabolism
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Phospholipid Transfer Proteins*
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Phospholipids / chemistry
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Phospholipids / metabolism*
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Proteolipids / chemistry
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Spectrometry, Fluorescence
Substances
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Carrier Proteins
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Detergents
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Fluorescent Dyes
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Lipid Bilayers
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Membrane Proteins
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Phosphatidylcholines
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Phosphatidylethanolamines
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Phospholipid Transfer Proteins
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Phospholipids
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Proteolipids
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proteoliposomes
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1,2-(palmitoyl-NBD-aminocaproyl)phosphatidylethanolamine
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1-palmitoyl-2-(6-(N-(7-nitrobenz)-2-oxa-1,3-diazol-4-yl)aminocaproyl)phosphatidylcholine
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Endopeptidase K
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4-Chloro-7-nitrobenzofurazan