LETHALITY OF THE COVALENT LINKAGE BETWEEN MISLOCALIZED MAJOR OUTER-MEMBRANE LIPOPROTEIN AND THE PEPTIDOGLYCAN OF ESCHERICHIA-COLI

The major outer membrane lipoprotein (Lpp) of Escherichia coli possess es serine at position 2, which is thought to function as the outer mem brane sorting signal, and lysine at the C terminus, through which Lpp covalently associates with peptidoglycan. Arginine (R) is present befo re the C-terminal lysine in the wild-type Lpp (LppSK). By replacing se rine (S) at position 2 with aspartate (D), the putative inner membrane sorting signal, and by deleting lysine (K) at the C terminus, Lpp mut ants with a different residue at either position 2 (LppDK) or the C te rminus (LppSR) or both (LppDR) were constructed, Expression of LppSR a nd LppDR little affected the growth of E, coli, In contrast, the numbe r of viable cells immediately decreased when LppDK was expressed, Prol onged expression of LppDK inhibited separation of the inner and outer membranes by sucrose density gradient centrifugation, whereas short-te rm expression did not, Pulse-labeled LppDK and LppDR were localized in the inner membrane, indicating that the amino acid residue at positio n 2 functions as a sorting signal for the membrane localization of Lpp , LppDK accumulated in the inner membrane covalently associated with t he peptidoglycan and thus prevented the separation of the two membrane s, Globomycin, an inhibitor of lipoprotein-specific signal peptidase I I, was lethal for E, coli only when Lpp possessed the C-terminal lysin e. Taken together, these results indicate that the inner membrane accu mulation of Lpp per se is not lethal for E, coli, Instead, a covalent linkage between the inner membrane Lpp having the C-terminal lysine an d the peptidoglycan is lethal for E, coli, presumably due to the disru ption of the cell surface integrity.