Distribution of murein‐lipoprotein between the cytoplasmic and outer membrane of <i>Escherichia coli</i>

Bosch, V.; Braun, Volkmar

doi:10.1016/0014-5793(73)80818-x

Cited by 55 publications

(30 citation statements)

References 9 publications

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“…most abundant proteins in the cell. It is also one of the Both forms are exclusively localized in the outer most extensively studied membrane proteins with membrane [4,5]. A more precise localization of the regard to its primary and secondary structures and lipoprotein molecules in the outer membrane is its biosynthesis (summarized in [l -31).…”

Section: Lipoprotein Treated With Boilingmentioning

confidence: 99%

Antigenic Determinants of Murein Lipoprotein and Its Exposure at the Surface of Enterobacteriaceae

Braun

Bosch

Klumpp

et al. 1976

European Journal of Biochemistry

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Murein lipoprotein from the outer membrane of Escherichia coli could be fixed to erythrocytes without pretreatment of the erythrocytes. Passive hemagglutination or immune hemolysis could thus be used as sensitive assays to determine antibodies against lipoprotein. In rabbit antisera prepared against whole cells of E. coli, Sulmonellu, Arizona and Shigellu antibodies against lipoprotein were present. The respective titers were lowest in encapsulated smooth strains and highest in rough mutants. Antisera against deep rough mutants showed even higher anti-lipoprotein titers than anti-R-lipopolysaccharide titers. Correspondingly, absorption of lipoprotein antibodies with enterobacterial strains was most pronounced with deep rough mutants and lowest with smooth strains. Lipoprotein becomes increasingly an immunogen as well as an antigen the more sugar residues are missing in the lipopolysaccharide on the cell surface. In wild-type cells lipoprotein is buried in the outer membrane; its exposure in mutant cells is related to defects at the cell surface.Antisera were prepared in rabbits against erythrocytes coated with various lipoprotein samples which differ at the C-terminal end of the polypeptide chain. Antibodies against lipoprotein containing two attached muropeptides (called lipoprotein I) are directed against the muropeptide attachment region. Antibodies against lipoprotein ending with lysine-55 (lipoprotein 11) were directed specifically against this terminal lysine residue. No serological reaction was observable after splitting off this lysine residue by carboxypeptidase B. Remarkably this antiserum against lipoprotein I1 neither reacted with the free lipoprotein ending with lysine-58 (lipoprotein 1V) nor with the muropeptide containing lipoprotein I. Antisera against the lipoprotein lacking lysine-55 (lipoprotein 111) reacted with all lipoprotein samples as did the antibodies in antisera prepared against whole heat-killed cells, thus indicating a common binding site located in the polypeptide chain towards the N-terminal end. No indication of an antigenic reactivity of the covalently linked lipoid was obtained.sodium dodecylsulfate and acetone reacted strongly with antisera against the native lipoprotein. The recovery of the antigenic sites demonstrates the remarkable stability of the native conformation.The lipoprotein is a new common antigen for closely related Enterobacteriaceae. Lipoprotein antisera might be useful for characterization of outer membrane mutants of Enterobacreriaceue.Lipoprotein treated with boiling 4

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Section: Lipoprotein Treated With Boilingmentioning

confidence: 99%

Antigenic Determinants of Murein Lipoprotein and Its Exposure at the Surface of Enterobacteriaceae

Braun

Bosch

Klumpp

et al. 1976

European Journal of Biochemistry

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“…One of the major outer membrane proteins in Gram-negative enteric bacteria is the murein lipoprotein discovered by Braun and his coworkers (1,2). More recently, Inouye and his coworkers (3) have characterized a precursor form of lipoprotein, the prolipoprotein, which contains 20 extra amino acids at the amino terminus.…”

mentioning

confidence: 99%

An Escherichia coli mutant with an amino acid alteration within the signal sequence of outer membrane prolipoprotein.

Lin¹,

Kanazawa²,

Ozols³

et al. 1978

Proc. Natl. Acad. Sci. U.S.A.

131

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Lipoprotein has been purified from an Escherichia coli strain carrying a mutation in the structural gene for murein lipoprotein (mlpA). Amino acid analysis of the purified mutant lipoprotein indicates that the mutant lipoprotein corresponds to the uncleaved prolipoprotein with a single amino acid replacement of glycine with aspartic acid. Automated Edman degradation has established the precise location of this amino acid substitution to be at the 14th residue of the prolipoprotein. This alteration in the signal sequence of prolipoprotein results in a failure of the mutated prolipoprotein to be jrocessed. Furthermore, the structural alteration in the mutant ipoprotein appears also to have affected its topological localization in the mutant cell. Whereas lipoprotein in the wild-type strain is exclusively located in the outer membrane of the cell envelope, the membrane-bound lipoprotein in this mutant is recovered in both the inner and outer membranes of the cell envelope. The data suggest, however, that proteolytic cleavage of prolipoprotein to form mature lipoprotein is not essential for the translocation and assembly of lipoprotein into the outer membrane.One of the major outer membrane proteins in Gram-negative enteric bacteria is the murein lipoprotein discovered by Braun and his coworkers (1, 2). More recently, Inouye and his coworkers (3) have characterized a precursor form of lipoprotein, the prolipoprotein, which contains 20 extra amino acids at the amino terminus. It has been suggested that these extra amino acids at the amino terminus of prolipoprotein may constitute the so-called signal sequence, postulated to play an important role in the biogenesis of this outer membrane protein.We have previously described the isolation and characterization of an Escherichia coli mutant altered in the structure of murein lipoprotein (4). The biochemical phenotype of this mutant lipoprotein includes a deficiency in covalently linked diglyceride, a defect in the assembly of the free form of mutant lipoprotein into the murein sacculus, and an apparent increase in the size of the mutant lipoprotein as compared to that of wild type (5). In this paper, we present evidence showing that the mutant lipoprotein corresponds to the uncleaved prolipoprotein with a single amino acid replacement within the signal sequence of prolipoprotein, a glycyl residue at position 14 being substituted by aspartate. We also find that while the majority of the mutant lipoprotein is assembled into the outer membrane of the cell envelope, there is a significant amount of lipoprotein in the inner membrane. MATERIALS AND METHODSBacterial Strains and Media. The E. coli strains used in the present study were wild-type E600 (mipA +) and mutant E602 (mipA -) as described (4). Isogenic transductant strains E613 (mipA + ) and E614 (mplA-) (6) were used in the study of lipoprotein localization in the cell envelope. Media used in the present study included M9 minimal medium and proteose peptone beef extract broth (6) Purification of Murein Lipoprote...

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“…We have shown that this lipoprotein exists in two different forms in the E. coli envelope (9, 10): a free form and a bound form, which is covalently linked to the peptidoglycan (11,12). There is twice as much of the free form as the bound form (9), and both forms are located exclusively in the outer membrane (13,14). The number of lipoprotein molecules per cell has been estimated to be 5 X 105 and 2.5 X 105 for the free and the bound forms, respectively (9,15).…”

mentioning

confidence: 99%

“…Assembly in the Outer Membrane. Since both the free and the bound forms of the lipoprotein are located exclusively ill the outer membrane (13,14), there are two possible ways for the present models to interact with the outer membrane: (i)…”

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confidence: 99%

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A Three-Dimensional Molecular Assembly Model of a Lipoprotein from the Escherichia coli Outer Membrane

Inouye

1974

Proc. Natl. Acad. Sci. U.S.A.

120

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From the complete amino-acid sequence of a lipoprotein from the outer membrane of E. coli, a three-dimensional molecular assembly model was constructed. It is proposed that the model provides a tubular hydrophilic channel through the outer membrane, which serves as a passive diffusion pore. An a-helix is constructed from the sequence, and six of them are arranged to form a superhelix with a hydrophilic interior and hydrophobic outer surface. The superhelical assembly is stabilized by seven ionic interactions between adjacent a-helices.Since the height of the assembly is 76 A, it could be inserted into the outer membrane and span the full 75-A thick membrane. The assembly is stabilized in the outer membrane not only by hydrophobic interaction between the surface of the assembly and the lipid bilayer, but also by three hydrocarbon chains of fatty acids linked to the amino-terminal end of the lipoprotein, which are flipped back along the assembly and inserted into the lipid bilayer of the outer membrane. Any two a-helices in an assembly are linked to the peptidoglycan at their carboxyl-terminal ends so that the outer membrane is anchored on the peptidoglycan layer. Six or more a-helices can form an assembly of this type. However, assuming that an assembly consists of six helices, there are 1.25 X 106 per cell hydrophilic channels of a diameter of 12.5 X and 35% of the cell surface is occupied by the assemblies.

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Distribution of murein‐lipoprotein between the cytoplasmic and outer membrane of Escherichia coli

Cited by 55 publications

References 9 publications

Antigenic Determinants of Murein Lipoprotein and Its Exposure at the Surface of Enterobacteriaceae

Antigenic Determinants of Murein Lipoprotein and Its Exposure at the Surface of Enterobacteriaceae

An Escherichia coli mutant with an amino acid alteration within the signal sequence of outer membrane prolipoprotein.

A Three-Dimensional Molecular Assembly Model of a Lipoprotein from the Escherichia coli Outer Membrane

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