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Lipoprotein prepared from the outer membrane of Salmonella typhimurium is a polyclonal activator of murine B-lymphocytes. It was shown to be mitogenic for splenic cultures, stimulating increased incorporation of [3H]thymidine into DNA. When injected intravenously into mice, the lipoprotein induced splenomegaly and polyclonal B-cell activation. The latter was evident from an increase in the number of plaque-forming cells against trinitrophenylated sheep erythrocytes. Similar results were obtained with Escherichia coli lipoprotein.

Protein I from the outer membrane of Escherichia coli and other members of the family Enterobacteriaceae is a potent mitogen and polyclonal B-lymphocyte activator. To determine the part of the polypeptide responsible for biological activity, we cleaved the molecule into defined polypeptide fragments of approximate molecular weights 24,000, 15,000, 9,000, 7,000, and 3,000 by using the cyanogen bromide method. The fragments were purified by gel permeation chromatography and by preparative polyacrylamide gel electrophoresis. They were investigated for mitogenicity and for the induction of immunoglobulin synthesis in lymphocyte cultures from several inbred mouse strains. The fragment of molecular weight 24,000 turned out to be a potent polyclonal B-lymphocyte activator comparable to native protein I. The low-molecular-weight fragments exhibited only marginal effects. Neither purified T lymphocytes nor thymocytes were activated. Our results show that a defined fragment of protein I is responsible for its lymphocyte-stimulating activity.

From the mutant bacterial strain Escherichia coli JE5511 lpp lpm, muropeptide-containing and muropeptide-free lipoproteins were prepared. By gas chromatography and by infrared spectroscopy we showed that the products were deficient in the two ester-bound N-terminal fatty acids, but still carried the amide-linked fatty acid. Mutant lipoproteins were tested for mitogenicity in lipopolysaccharide nonresponder C3H/HeJ mice by incorporation of [3H]thymidine and [3H]uridine and by hemolytic plaque assays for immunoglobulin-secreting plasma cells. Our results showed that the mutant lipoproteins still exhibited marked mitogenicity toward mouse B-lymphocytes, although the activity of the products was reduced in comparison to the wild-type lipoprotein. Thus, the presence of one fatty acid in the N-terminal part of lipoprotein is sufficient to bring about mitogenicity.