Expression of human IL-1 beta in Salmonella typhimurium. A model system for the delivery of recombinant therapeutic proteins in vivo.

The feasibility of using Salmonella typhimurium aroA mutant (SL3261) to deliver protein therapeutic agents was investigated in a murine model system. We have constructed an Escherichia coli expression plasmid designed to express the human protein IL-1 beta. This plasmid expresses IL-1 beta to high levels (greater than 30% total cell protein) in E. coli. In Salmonella the IL-1 beta is expressed constitutively to about 10% total cell protein, as verified by Western blotting analysis using polyclonal rabbit anti-IL-1 beta antibody. The protein is produced in a soluble and biologically active form. BALB/c mice administered orally or i.v. with S. typhimurium aroA mutants carrying the plasmid produced highly significant antibody responses against human IL-1 beta as determined by a solid-phase RIA. Furthermore, mice injected with the construct were significantly protected against lethal gamma-irradiation (850 rad). This study therefore demonstrates that the vaccine strain of Salmonella mutants can also be used effectively to deliver therapeutic proteins in vivo.

Analysis of the chain length of oligomers and polymers of sialic acid isolated from Neisseria meningitidis group B and C and Escherichia coli K1 and K92.

A series of (2----8)-alpha-, (2----9)-alpha-, and alternate (2----8)-alpha- and (2----9)-alpha-linked oligomers of sialic acid (N-acetylneuraminic acid, NeuNAc) was prepared by digestion with bacteriophage or by partial hydrolysis at pH 7.0 and 100 degrees of polymers of sialic acid produced by Neisseria meningitidis and Escherichia coli. The oligosaccharides were purified by gel filtration or by anion-exchange chromatography, and their chain lengths were determined by colorimetric measurement of the formaldehyde released from the non-reducing end residue after periodate oxidation, radiolabelling of the reducing end residue by reduction with borotritiide, and determination of the ratio of the non-reducing end and internal residues by g.l.c. of the trimethylsilyl derivatives of the methyl ester methyl beta-ketosides. 1H-N.m.r. spectroscopy was used to confirm the chain length of two oligosaccharides. These methods were used to determine the average chain-length of the sialic acid polysaccharides produced by N. meningitidis and E. coli and the percentage of chains with covalently bound lipid moieties at the reducing end.