The deletion of 70 amino acids near the N-terminal end of the sucrose-specific porin ScrY causes its functional similarity to LamB in vivo and in vitro.
A deletion mutant ScrY delta 3-73 of the sucrose-specific porin ScrY was constructed in which 70 amino acids of the mature protein were deleted near the N-terminal end. ScrY delta 3-72 was still able to oligomerize and inserted properly into the outer membrane of an Escherichia coli strain. The protein was isolated and purified by standard procedures. The mutant protein showed, in contrast to wild-type ScrY, a tight association with the murein. Reconstitution experiments with artificial lipid bilayer membranes demonstrated that ScrY delta 3-72 produced defined cation-selective channels in planar lipid bilayers. Its single-channel conductance was reduced to about half of the value of wild-type ScrY. The deletion had a relatively small influence on the stability constants for carbohydrate binding. However, in contrast to wild-type ScrY, [14C]-maltopentaose was efficiently taken up into whole E. coli cells containing ScrY delta 3-72. The sequence of the N-terminus of mature ScrY was identified as starting with glutamine 23. The possible structure of ScrY and ScrY delta 3-72 in the outer membrane is discussed.
Schülein
K
K
Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum) der Universität Würzburg, Germany.
Andersen
C
C
Benz
R
R
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D001692:Biological Transport; D050260:Carbohydrate Metabolism; D002412:Cations / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D008051:Lipid Bilayers / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009844:Oligosaccharides / Q000378:metabolism; D018272:Porins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017433:Protein Structure, Secondary; D011991:Receptors, Virus / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D017421:Sequence Analysis; D017384:Sequence Deletion; D013395:Sucrose / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002412:Cations; D029968:Escherichia coli Proteins; D008051:Lipid Bilayers; D009844:Oligosaccharides; D018272:Porins; D011991:Receptors, Virus; D011994:Recombinant Proteins; C489789:ScrY protein, E coli; C079242:ScrY protein, bacteria; C016532:maltoporins; C019193:maltopentaose; D013395:Sucrose