Novel bacterial lipoprotein structures conserved in low-GC content gram-positive bacteria are recognized by Toll-like receptor 2.
Bacterial lipoproteins/lipopeptides inducing host innate immune responses are sensed by mammalian Toll-like receptor 2 (TLR2). These bacterial lipoproteins are structurally divided into two groups, diacylated or triacylated lipoproteins, by the absence or presence of an amide-linked fatty acid. The presence of diacylated lipoproteins has been predicted in low-GC content gram-positive bacteria and mycoplasmas based on the absence of one modification enzyme in their genomes; however, we recently determined triacylated structures in low-GC gram-positive Staphylococcus aureus, raising questions about the actual lipoprotein structure in other low-GC content gram-positive bacteria. Here, through intensive MS analyses, we identified a novel and unique bacterial lipoprotein structure containing an N-acyl-S-monoacyl-glyceryl-cysteine (named the lyso structure) from low-GC gram-positive Enterococcus faecalis, Bacillus cereus, Streptococcus sanguinis, and Lactobacillus bulgaricus. Two of the purified native lyso-form lipoproteins induced proinflammatory cytokine production from mice macrophages in a TLR2-dependent and TLR1-independent manner but with a different dependence on TLR6. Additionally, two other new lipoprotein structures were identified. One is the "N-acetyl" lipoprotein structure containing N-acetyl-S-diacyl-glyceryl-cysteine, which was found in five gram-positive bacteria, including Bacillus subtilis. The N-acetyl lipoproteins induced the proinflammatory cytokines through the TLR2/6 heterodimer. The other was identified in a mycoplasma strain and is an unusual diacyl lipoprotein structure containing two amino acids before the lipid-modified cysteine residue. Taken together, our results suggest the existence of novel TLR2-stimulating lyso and N-acetyl forms of lipoproteins that are conserved in low-GC content gram-positive bacteria and provide clear evidence for the presence of yet to be identified key enzymes involved in the bacterial lipoprotein biosynthesis.
Global Research Laboratory of Insect Symbiosis, College of Pharmacy, Pusan National University, Busan 609-735, Korea.
Kenji
K
Kurokawa
Kyoung-Hwa
KH
Ryu
Rie
R
Ichikawa
Akiko
A
Masuda
Min-Su
MS
Kim
Hanna
H
Lee
Jun-Ho
JH
Chae
Takashi
T
Shimizu
Tatsuya
T
Saitoh
Koichi
K
Kuwano
Shizuo
S
Akira
Naoshi
N
Dohmae
Hiroshi
H
Nakayama
Bok Luel
BL
Lee
D000818:Animals; D001409:Bacillus cereus / Q000276:immunology / Q000378:metabolism; D001412:Bacillus subtilis / Q000276:immunology / Q000378:metabolism; D013293:Enterococcus faecalis / Q000276:immunology / Q000378:metabolism; D056506:Geobacillus / Q000276:immunology / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000276:immunology* / Q000378:metabolism; D007778:Lactobacillus / Q000276:immunology / Q000378:metabolism; D017737:Macrophages, Peritoneal / Q000276:immunology* / Q000382:microbiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D011019:Pneumonia, Mycoplasma / Q000276:immunology / Q000378:metabolism; D013298:Streptococcus sanguis / Q000276:immunology / Q000378:metabolism; D051194:Toll-Like Receptor 1 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D051195:Toll-Like Receptor 2 / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D051216:Toll-Like Receptor 6 / Q000235:genetics / Q000276:immunology / Q000378:metabolism
C493486:Tlr2 protein, mouse; C495369:Tlr6 protein, mouse; D051194:Toll-Like Receptor 1; D051195:Toll-Like Receptor 2; D051216:Toll-Like Receptor 6