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The amino terminal region of escherichia coli lactose permease mediates membrane contact of the nascent polypeptide chain






European Journal of Biochemistry 163(3): 653-658

The amino terminal region of escherichia coli lactose permease mediates membrane contact of the nascent polypeptide chain

Plasmids encoding N-terminal segments of the Escherichia coli lactose permease (also referred to as lactose carrier) have been used to analyze the biosynthesis and membrane insertion of this complex integral protein of the cytoplasmic membrane. Such truncated polypeptides were found to be stably associated with the membrane and to resemble the full-length protein with respect to their solubilization characteristics. Membrane-bound and free cytoplasmic polysomes were prepared from plasmid-bearing cells and incubated in the presence of [35S]methionine to permit completion of polypeptides initiated in vivo. Under these conditions, lactose permease was found to be radiolabeled in the fraction of membrane-bound polysomes; .beta.-galactosidase, used as a control, was translated almost exclusively by free polysomes. From similar experiments with N-terminal segments of lactose permease, we estimate that at most a polypeptide of 120 amino acid residues emerging from the ribosome is needed to target the nascent chain to the lipid bilayer and to mediate attachment of the ribosome to the membrane during elongation. Additional data support the idea that even shorter N-terminal sequences of 50 and 71 amino acid residues contain sufficient 'information' to provide contact with the membrane.


Accession: 006599481



Related references

Stochaj, U.; Ehring, R., 1987: The N-terminal region of Escherichia coli lactose permease mediates membrane contact of the nascent polypeptide chain. Plasmids encoding N-terminal segments of the Escherichia coli lactose permease (also referred to as lactose carrier) have been used to analyze the biosynthesis and membrane insertion of this complex integral protein of the cytoplasmic membrane. Su...

MacIntyre, S.; Eschbach, M.L.; Schwarz, H.; Ehring, R., 1989: Topological analysis of the amino-terminal region of lactose permease using the Escherichia coli outer membrane protein, OmpA, as a marker. LacY-ompA fusions, encoding the N-terminal 50, 71 or 143 residues of lactose permease, were constructed. The observed orientation of the OmpA part of each hybrid protein with respect to the plasma membrane supports current models of the N-terminus...

Bibi, E.; Stearns, S.M.; Kaback, H.R., 1992: The N-terminal 22 amino acid residues in the lactose permease of Escherichia coli are not obligatory for membrane insertion or transport activity. When the lactose (lac) permease of Escherichia coli is expressed from the lac promoter at relatively low rates, deletion of amino acid residues 2-8 (delta 7) or 2-9 (delta 8) from the hydrophilic N terminus has a relatively minor effect on the abi...

Bibi, E.; Stearns, S.M.; Kaback, H.Ronald, 1992: The N-terminal 22 amino acid residues in the lactose permease of Escherichia coli are not obligatory for membrane insertion of transport activity. Proceedings of the National Academy of Sciences of the United States of America 89: 80-4

Macintyre S.; Eschbach M L.; Schwartz H.; Ehring R., 1989: Topological analysis of the amino terminal region of lactase permease using the escherichia coli outer membrane protein omp a as a marker. Febs Letters 247(2): 396-400

Sahin-Tóth, M.; Persson, B.; Schwieger, J.; Cohan, P.; Kaback, H.R., 1994: Cysteine scanning mutagenesis of the N-terminal 32 amino acid residues in the lactose permease of Escherichia coli. Using a functional lactose permease mutant devoid of Cys residues (C-less permease), each amino acid residue in the hydrophilic N-terminus and the first putative transmembrane helix was systematically replaced with Cys (from Tyr-2 to Trp-33). Twen...

Yamada Y.; Chang Y Y.; Daniels G.A.; W.L.F.; Tomich J.M.; Yamada M.; Saier M.H.Jr, 1991: Insertion of the mannitol permease into the membrane of escherichia coli possible involvement of an amino terminal amphiphilic sequence. The in vivo membrane assembly of the mannitol permease, the mannitol Enzyme II (IImtl) of the Escherichia coli phosphotransferase system, has been studied employing molecular genetic approaches. Removal of the N-terminal amphiphilic leader of the...

Wu, J.; Sun, J.; Kaback, H.R., 1996: Purification and functional characterization of the C-terminal half of the lactose permease of Escherichia coli. The lactose permease has been expressed in contiguous, non-overlapping polypeptide fragments containing the N-terminal (N6) and C-terminal (C6) transmembrane domains of the protein [Bibi, E., & Kaback, H. R. (1990) Proc. Natl. Acad. Sci. U.S.A...

Yamato, I., 1992: Membrane assembly of lactose permease of Escherichia coli. Lactose permease, the lacY gene product in Escherichia coli, is an integral membrane protein. Its induction was examined in secAts and secYts mutants by measuring o-nitrophenyl-.beta.-galactoside uptake activity. In contrast to the synthesis of th...

Kaback, H.R., 2000: The lactose permease of Escherichia coli What to do while awaiting crystals of a membrane protein and thereafter. Biophysical Journal. 78(1 Part 2): 12a,.