title: Insertion and folding of the amino-terminal amphiphilic signal sequences of the mannitol and glucitol permeases of Escherichia coli.
abstract: Peptides which correspond to the NH2-terminal 23 or 22 residues of the mannitol and glucitol permeases (enzymes IImtl and IIgut of the bacterial phosphotransferase system; mtl-23 and gut-22) and which are believed to function in envelope targeting were synthesized chemically, and their interactions with lipid model membranes were studied. Both wild-type peptides penetrated phospholipid monolayers up to high surface pressures, and partition constants of 8.0 x 10(4) M-1 and 4.2 x 10(4) M-1, respectively, were derived from the incorporation isotherms of mtl-23 and gut-22 with monolayers of 1-palmitoyl-2-oleoyl-3-sn-phosphatidylcholine at 32 mN/m or bilayers of the same lipid. The mtl-23 peptide was highly alpha-helical in trifluoroethanol, sodium dodecyl sulfate, lysolecithin, or vesicles of 1-palmitoyl-2-oleoyl-3-sn-phosphatidylglycerol, with estimated percentages of alpha-helix ranging between 60 and 85%. The interactions with model membranes of several single site mutants (S3P, D4P, and D4K) of mtl-23 which were defective in properly assembling the mannitol permease in the cytoplasmic membrane of Escherichia coli were also studied. The contents of alpha-helix of these peptides in detergent micelles or phospholipid bilayers were not significantly changed compared with those of the wild type, suggesting that the amphiphilic NH2-terminal membrane-targeting domain could still be formed in these mutants. However, the mutants which contained a proline in positions 3 or 4, i.e. NH2-terminal to the proposed amphiphilic alpha-helix, partitioned into phospholipid monolayers with partition constants that were 2 or 4 times smaller than those of the wild type. Based on these data, a model of the amphiphilic structure of the NH2-terminal domain of the mannitol permease is discussed. This domain may interact physiologically with amphiphilic interfaces of lipids and/or proteins during membrane insertion.
authors.affiliation: Department of Physiology, University of Virginia School of Medicine, Charlottesville 22908.
authors.forename: S H
authors.initials: SH
authors.lastname: Portlock
authors.forename: Y
authors.initials: Y
authors.lastname: Lee
authors.forename: J M
authors.initials: JM
authors.lastname: Tomich
authors.forename: L K
authors.initials: LK
authors.lastname: Tamm
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism; D029968:Escherichia coli Proteins; D008051:Lipid Bilayers; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D021382:Protein Sorting Signals / Q000235:genetics*; D013816:Thermodynamics
chemical_list: D029968:Escherichia coli Proteins; D008051:Lipid Bilayers; D009004:Monosaccharide Transport Proteins; D021382:Protein Sorting Signals; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C022358:mannitol PTS permease, E coli; C058558:phosphoenolpyruvate glucitol phosphotransferase