Glycine-144 is required for efficient folding of outer membrane protein PhoE of Escherichia coli K12

Abstract

Short stretches of similar sequences have been detected in unrelated bacterial outer membrane proteins (Nikaido and Wu (1984) Proc. Natl. Acad. Sci. USA 81, 1048-1052). In the most pronounced similarity region, only a glycine residue is absolutely conserved. To investigate whether this glycine residue is essential for outer membrane incorporation, oligonucleotide-directed mutagenesis was applied to replace this residue, i.e. Gly-144, as well as two other Gly-residues in pore protein PhoE. Substitution of Gly-52 and Gly-258 by Ala And Val, respectively, did not influence outer membrane incorporation. However, the substitution of Gly-144 by Leu affected the efficiency of outer membrane incorporation. After in vitro synthesis this mutant protein was less efficiency precipitated with monoclonal antibodies that recognize coformational epitopes than wild-type PhoE, showing that the mutation interferes with correct folding of the protein.