Beta-barrels covalently link peptidoglycan and the outer membrane in the alpha-proteobacterium Brucella abortus

Gram negative bacteria are surrounded by a cell envelope that comprises an outer membrane (OM) and an inner membrane (IM) which together delimit the periplasmic space, which contains peptidoglycan (PG). Covalent anchoring of the OM to the peptidoglycan is crucial for envelope integrity in Escherichia coli. When OM is not attached to PG the OM forms blebs and detaches from the cell. The Braun lipoprotein (Lpp) covalently attaches OM to PG but is only present in a small number of gamma-proteobacteria, which leaves the mechanism of OM-PG attachment in other species undetermined. We report that the OM is attached to the peptidoglycan by covalent crosslinks between the N-termini of integral OM beta-barrel-shaped proteins (OMPs) and the peptide stems of PG in the alpha-proteobacteria Brucella abortus and Agrobacterium tumefaciens. Crosslinking is catalyzed by L,D-transpeptidases and  attached OMPs have a conserved alanyl-aspartyl motif at their N-terminus. Mutation of the aspartate in this motif prevents OMP crosslinking and results in OM membrane instability. The alanyl-aspartyl motif  is conserved in OMPs from Rhizobiales, which means that it is feasible that OMP-PG crosslinks are widespread in alpha-proteobacteria