Role of the Arg¹⁵⁸ residue of the outer membrane PhoE pore protein of Escherichia coli K 12 in bacteriophage TC 45 recognition and in channel characteristics

Abstract

(Rcccived Novcinher 29. 1 9 W M a r c h 5. 1985) --EJB 841257 In order t o study the structure-function relationship o f t h e PhoE protein pore we have isolated five independent. 'I'C'45-resistant, plzoE mutants all of which appeared to produce normal amounts of a n clectrophoretically altered PhoE protein, designated a s PhoE * protein. Nticleotidc sequence analysis of the DNA liagments carrying the mutations showed that the mutations all correspond to a G C to A . 'r transition at the same place within the phoE gene resulting in a deduced change of amino acid residue arginine 158 into histidine. 'This result shows that the arginine 158 residue plays ;in important role in thc interaction of the PhoE protein pore with phage TC45.Moloreover, studies on the channel properties o!' the PhoE * protein showed that the PhoE channel has lost part of its preference for negatively charged solutes. ;IS a result of the arginine to histidine changc. 'l'he results are discussed in terms of the structure-function relationship of PhoE protein ;is well as in terms o f thc topological organization of the protein channel in the outer membrane.

Voltage sensing in the PhoE and OmpF outer membrane porins of Escherichia coli: role of charged residues

“…Analysis of whole-cell protein patterns by SDS/ polyacrylamide gel electrophoresis (Lugtenberg et al, 1975) revealed that the various PhoE mutant proteins were equally well expressed as the wildtype protein after overnight growth of plasmidcontaining derivatives of strain CE1265 (Korteland et al, 1985) in L-broth (Tommassen et al, 1983; results not shown). In a whole-cell enzyme-linked immunosorbent assay (van der Ley et al, 1985), all mutants were recognized by PP4-1, a monoclonal antibody that recognizes a conformational, cell-surface-exposed epitope of the PhoE protein.…”

Efficient recognition by rat T cell clones of an epitope of mycobacterial hsp 65 inserted in Escherichia coli outer membrane protein PhoE

“…coli K 12 strain CE1224 [ 171 produces no pore proteins due to mutations in the phoE and ompR genes. E. coli strain CE1248 [18] also containsphoE and ompR mutations and, in addition, a phoR mutation resulting in constitutive expression of the pho regulon. Cells were grown overnight at 37°C under aeration in L broth [17] or in a phosphatelimited medium described by Levinthal et al [19].…”

A colorimetric detection system for Calymmatobacterium granulomatis

“…8 Korteland et al reported that all E coli K 12 strains tested in their laboratory with resistance to the PhoE specific bacteriophage TC45 showed a mutation at Arg158. 9 While unproved, it is plausible that the C granulomatis isolates present in Australia have also acquired this selective advantage at some point in their evolution.…”