Interaction of the transmembrane domain of lysis protein E from bacteriophage ϕX174 with bacterial translocase MraY and peptidyl-prolyl isomerase SlyD

Abstract

The molecular target for the bacteriolytic E protein from bacteriophageϕX174, responsible for host cell lysis, is known to be the enzyme phospho-MurNAc-pentapeptide translocase (MraY), an integral membrane protein involved in bacterial cell wall peptidoglycan biosynthesis, with an essential role being played by peptidyl-prolyl isomerase SlyD. A synthetic 37 aa peptide Epep, containing the N-terminal transmembraneα-helix of E, was found to be bacteriolytic againstBacillus licheniformis, and inhibited membrane-bound MraY. The solution conformation of Epepwas found by circular dichroism (CD) spectroscopy to be 100 %α-helical. No change in the CD spectrum was observed upon addition of purifiedEscherichia coliSlyD, implying that SlyD does not catalyse prolyl isomerization upon E. However, Epepwas found to be a potent inhibitor of SlyD-catalysed peptidyl-prolyl isomerization (IC500.15 μM), implying a strong interaction between E and SlyD. Epepwas found to inhibitE. coliMraY activity when assayed in membranes (IC500.8 μM); however, no inhibition of solubilized MraY was observed, unlike nucleoside natural product inhibitor tunicamycin. These results imply that the interaction of E with MraY is not at the MraY active site, and suggest that a protein–protein interaction is formed between E and MraY at a site within the transmembrane region.