Dissemination of lipid A deacylases (pagL) among gram-negative bacteria: identification of active-site histidine and serine residues.
Lipopolysaccharide (LPS) is one of the main constituents of the Gram-negative bacterial outer membrane. It usually consists of a highly variable O-antigen, a less variable core oligosaccharide, and a highly conserved lipid moiety, designated lipid A. Several bacteria are capable of modifying their lipid A architecture in response to external stimuli. The outer membrane-localized lipid A 3-O-deacylase, encoded by the pagL gene of Salmonella enterica serovar Typhimurium, removes the fatty acyl chain from the 3 position of lipid A. Although a similar activity was reported in some other Gram-negative bacteria, the corresponding genes could not be identified. Here, we describe the presence of pagL homologs in a variety of Gram-negative bacteria. Although the overall sequence similarity is rather low, a conserved domain could be distinguished in the C-terminal region. The activity of the Pseudomonas aeruginosa and Bordetella bronchiseptica pagL homologs was confirmed upon expression in Escherichia coli, which resulted in the removal of an R-3-hydroxymyristoyl group from lipid A. Upon deacylation by PagL, E. coli lipid A underwent another modification, which was the result of the activity of the endogenous palmitoyl transferase PagP. Furthermore, we identified a conserved histidine-serine couple as active site residues, suggesting a catalytic mechanism similar to serine hydrolases. The biological function of PagL remains unclear. However, because PagL homologs were found in both pathogenic and nonpathogenic species, PagL-mediated deacylation of lipid A probably does not have a dedicated role in pathogenicity.
Geurtsen
Jeroen
J
Department of Molecular Microbiology, Utrecht University, Padualaan 8, Utrecht 3584 CH, The Netherlands. j.j.g.geurtsen@bio.uu.nl
Steeghs
Liana
L
Hove
Jan Ten
JT
van der Ley
Peter
P
Tommassen
Jan
J
D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D016950:Bordetella bronchiseptica / Q000378:metabolism; D002240:Carbohydrate Sequence; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry* / Q000378:metabolism; D002498:Centrifugation; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry; D004187:Disaccharides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D005998:Glycine / Q000031:analogs & derivatives* / Q000737:chemistry; D006639:Histidine / Q000737:chemistry*; D015151:Immunoblotting; D008050:Lipid A / Q000737:chemistry; D008055:Lipids / Q000737:chemistry; D008956:Models, Chemical; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000378:metabolism; D019779:Salmonella enterica / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization
D000906:Antibodies; D001426:Bacterial Proteins; D004187:Disaccharides; D008050:Lipid A; D008055:Lipids; D012694:Serine; D006639:Histidine; D004247:DNA; D002265:Carboxylic Ester Hydrolases; C424702:PagL protein, Salmonella typhimurium; D005998:Glycine; C100184:tricine