Post-translational modification and processing of Escherichia coli prolipoprotein in vitro.
Escherichia coli strain MM18 cells containing malE-lacZ hybrid protein was reported to accumulate prolipoprotein when they were induced with maltose [Ito, K., Bassford, P. J. & Beckwith, J. (1981) Cell 24, 707-717]. We have shown that the prolipoprotein accumulated in maltose-induced MM18 cells is not modified, lacking covalently linked glyceride. When the cell envelope of MM18 containing unmodified prolipoprotein was incubated in the presence of detergent with [2-3H]glycerol-labeled cell envelope of strain JE5505 lacking murein lipoprotein, incorporation of [2-3H]glycerol radioactivity into both prolipoprotein and processed mature lipoprotein was observed. Likewise, when [3H]-palmitate-labeled JE5505 cell envelope was incubated with the MM18 cell envelope containing unmodified prolipoprotein in the presence of detergent, [3H]palmitate radioactivity was incorporated into prolipoprotein by ester linkage and into mature lipoprotein by both ester and amide linkages. These results indicate that our in vitro system contains activities of prolipoprotein modification and processing enzymes, including glyceryltransferase, O-acyltransferase, signal peptidase, and N-acyltransferase. The signal peptidase activity in our in vitro system was completely inhibited by globomycin. At pH 5.0, glyceryltransferase was inactive. Signal peptidase was active at pH 5.0, provided that prolipoprotein had been modified by glyceryltransferase (O-acyl-transferase) during a prior incubation at pH 9.1. These results strongly suggest that the modification of prolipoprotein by glyceryltransferase (and O-acyltransferase) precedes, and may in fact be a prerequisite for, the processing of prolipoprotein by signal peptidase.
D001426:Bacterial Proteins / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D004843:Epistasis, Genetic; D004926:Escherichia coli / Q000235:genetics*; D005990:Glycerol / Q000378:metabolism; D066298:In Vitro Techniques; D008074:Lipoproteins / Q000235:genetics* / Q000378:metabolism; D010168:Palmitates / Q000378:metabolism; D011498:Protein Precursors / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D008074:Lipoproteins; D010168:Palmitates; D011498:Protein Precursors; D005990:Glycerol