A genetic approach for analyzing the pathway of LamB assembly into the outer membrane of Escherichia coli.
Results presented in this study demonstrate that a mutation which inserts an additional tyrosine between the 2 tyrosines at residues 118 and 119 of mature LamB protein results in a temperature-dependent assembly defect. This defect leads to the accumulation of an intermediate at the restrictive temperature that is most likely an assembly-defective monomer. These monomers are rapidly degraded in the wild type (htrA+) strain, and the biphasic kinetics of this degradation indicate that the mutation affects the assembly process and not the final product, i.e. stable trimers. In addition, our data show that the temperature-dependent assembly defect in the mutant strain is reversible, and therefore the accumulated monomers represent a true assembly intermediate. Fractionation studies show that the monomers, which can be accumulated in htrA (degP) mutants at the restrictive temperature, are associated with the outer membrane, indicating that trimerization of LamB is not a prerequisite for localization.
Misra
R
R
Department of Molecular Biology, Lewis Thomas Laboratory, Princeton University, New Jersey 08544-1014.
Peterson
A
A
Ferenci
T
T
Silhavy
T J
TJ
D018528:ATP-Binding Cassette Transporters; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002451:Cell Compartmentation; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000648:ultrastructure; D029968:Escherichia coli Proteins; D046911:Macromolecular Substances; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D018272:Porins; D011991:Receptors, Virus / Q000235:genetics* / Q000378:metabolism; D013329:Structure-Activity Relationship; D013696:Temperature
D018528:ATP-Binding Cassette Transporters; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D046911:Macromolecular Substances; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D018272:Porins; D011991:Receptors, Virus; C016532:maltoporins; C461599:maltose transport system, E coli