Isolation, structure elucidation, and synthesis of a macrophage stimulatory lipopeptide from Mycoplasma fermentans acting at picomolar concentration.
Macrophages are typically stimulated by components of microbial cell walls. Surprisingly, cell wall-less mycoplasmas can also very efficiently stimulate macrophages. We showed recently that mycoplasma-derived lipopeptides constitute the active principle. We have now isolated a clone of Mycoplasma fermentans expressing mainly one macrophage-stimulating lipopeptide. This lipopeptide was detergent-extracted and isolated by reversed-phase high-performance liquid chromotography, using nitric oxide release from C3H/HeJ mouse macrophages as bioassay for detection. In contrast to "conventional" bacterial lipoproteins, this lipopeptide had a free NH2 terminus. Amino acid composition, sequence, and the molecular weight of 2,163. 3 are consistent with the following structure: S-(2, 3-bisacyloxypropyl)cysteine-GNNDESNISFKEK with one mole C16:0, and a further mole of a mixture of C18:0 and C18:1 fatty acid per lipopeptide molecule. The sequence could not be found in either the protein identification resource nor the Swiss Prot data bank. We named this 2-kD lipopeptide, macrophage-activating lipopeptide-2 (MALP-2). Synthetic dipalmitoyl MALP-2 and mycoplasma-derived MALP-2 were compared with the bioassay. Both lipopeptides showed an identical dose dependency with a half-maximal response at 10(-11) M concentration. MALP-2 may be one of the most potent natural macrophage stimulators besides endotoxin.
Mühlradt
P F
PF
Immunobiology and Structure Research Groups, Gesellschaft für Biotechnologische Forschung mbH, D-38124 Braunschweig, Germany.
Kiess
M
M
Meyer
H
H
Süssmuth
R
R
Jung
G
G
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D055666:Lipopeptides; D008262:Macrophage Activation; D017737:Macrophages, Peritoneal / Q000276:immunology; D051379:Mice; D008809:Mice, Inbred C3H; D008970:Molecular Weight; D016858:Mycoplasma fermentans / Q000737:chemistry*; D009569:Nitric Oxide / Q000378:metabolism; D009842:Oligopeptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000596:Amino Acids; D055666:Lipopeptides; D009842:Oligopeptides; D009569:Nitric Oxide; C106335:macrophage stimulatory lipopeptide 2