An Escherichia coli mutant lacking the cold shock-induced palmitoleoyltransferase of lipid A biosynthesis: absence of unsaturated acyl chains and antibiotic hypersensitivity at 12 degrees C.
An acyltransferase induced by cold shock in Escherichia coli, designated LpxP, incorporates a palmitoleoyl moiety into nascent lipid A in place of the secondary laurate chain normally added by LpxL(HtrB) (Carty, S. M., Sreekumar, K. R., and Raetz, C. R. H. (1999) J. Biol. Chem. 274, 9677-9685). To determine whether the palmitoleoyl residue alters the properties of the outer membrane and imparts physiological benefits at low growth temperatures, we constructed a chromosomal insertion mutation in lpxP, the structural gene for the transferase. Membranes from the lpxP mutant MKV11 grown at 12 degrees C lacked the cold-induced palmitoleoyltransferase present in membranes of cold-shocked wild type cells but retained normal levels of the constitutive lauroyltransferase encoded by lpxL. When examined by mass spectrometry, about two-thirds of the lipid A molecules isolated from wild type E. coli grown at 12 degrees C contained palmitoleate in place of laurate, whereas the lipid A of cold-adapted MKV11 contained only laurate in amounts comparable with those seen in wild type cells grown at 30 degrees C or above. To probe the integrity of the outer membrane, MKV11 and an isogenic wild type strain were grown at 30 or 12 degrees C and then tested for their susceptibility to antibiotics. MKV11 exhibited a 10-fold increase in sensitivity to rifampicin and vancomycin at 12 degrees C compared with wild type cells but showed identical resistance when grown at 30 degrees C. We suggest that the palmitoleoyltransferase may confer a selective advantage upon E. coli cells growing at lower temperatures by making the outer membrane a more effective barrier to harmful chemicals.
Vorachek-Warren
Mara K
MK
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
Carty
Sherry M
SM
Lin
Shanhua
S
Cotter
Robert J
RJ
Raetz
Christian R H
CR
D000217:Acyltransferases / Q000378:metabolism*; D002240:Carbohydrate Sequence; D002462:Cell Membrane; D003080:Cold Temperature; D004247:DNA / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D008050:Lipid A / Q000096:biosynthesis*; D008055:Lipids; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012293:Rifampin / Q000494:pharmacology; D013696:Temperature; D013997:Time Factors; D014640:Vancomycin / Q000494:pharmacology
D029968:Escherichia coli Proteins; D008050:Lipid A; D008055:Lipids; D014640:Vancomycin; D004247:DNA; D000217:Acyltransferases; C118452:lpxP protein, E coli; D012293:Rifampin