Further genetic analysis of the C-terminal external loop region in Escherichia coli maltoporin.
LamB specifically facilitates the diffusion of maltose and maltodextrins through the bacterial outer membrane, and acts as a general (i.e. non-specific) porin for small hydrophilic molecules (< 600 daltons). We reported previously that deletion of the last predicted external domain near the C-terminus of the Eschirichia coli LamB protein (residues 376 to 405), affected in vivo the binding and transport of maltodextrins (specific pore functions), and also increased bacterial sensitivity to large antibiotics. The residues covered by this deletion correspond almost exactly to the major cell surface loop of LamB on the structural model based on X-ray crystallography (loop L9, residues 375 to 405). The L9 loop comprises a large central portion, which varies in size and sequence between the LamB proteins from different species. This variable region is flanked by two highly charged and conserved portions, which overlap with the adjacent beta strands. To identify subregions in L9 that influence the pore properties of LamB, we constructed and analysed nine mutants in loop L9 and its flanking sequences. Deletion of the 23-amino-acids central variable portion of the loop (residues 379 to 401), and deletion of the downstream conserved region (residues 402 to 409), only moderately affected specific maltoporin function. In contrast, deletion of the conserved region (residues 372 to 378) upstream of the variable portion strongly decreased specific maltoporin function and also increased sensitivity to large antibiotics, accounting for most, if not all, of the effects of the complete deletion of L9.
Department of Chemistry & Biochemistry, University of Oklahoma, Norman 73019, USA.
P E
PE
Klebba
S M
SM
Newton
A
A
Charbit
V
V
Michel
D
D
Perrin
M
M
Hofnung
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001425:Bacterial Outer Membrane Proteins; D010582:Bacteriophage lambda; D017124:Conserved Sequence; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D007700:Kinetics; D008320:Maltose / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D010641:Phenotype; D018272:Porins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011487:Protein Conformation; D011991:Receptors, Virus / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid
D001425:Bacterial Outer Membrane Proteins; D018272:Porins; D011991:Receptors, Virus; C016532:maltoporins; D008320:Maltose