Mapping and disruption of the cybB gene coding for cytochrome b561 in Escherichia coli

Abstract The cybB gene on a plasmid encoding cytochrome b ₅₆₁ in Escherichia coli was disrupted by insertion of Km^rl determinant DNA. The cromosomal cybB gene was replaced by the inactivated cybB gene on the plasmid by homologous recombination using λ phage lysogenization and heat-induction. The replacement was confirmed by Southern and Western blotting analyses. Deficiency on the cybB gene product did not affect the growth properties of the cells, and the oxidase activities of the cells dependent on various substrates were similar to those of the parental strain. Cytochrome b ₅₆₁ is concluded to be expressed in E. coli , but may not play a major role in cell growth. In the genetic map of E. coli , the cybB gene was determined by conjugational and transductional crosses to be at 31 min between trg and terC .     

Participation of hup gene product in ori2-dependent replication of fertility plasmid F

The fertility plasmid F'gal was not stably maintained in a hupA-hupB double mutant of Escherichia coli. Moreover, mini-F plasmids pFZY1, pFTC1 and pFTC2 were unable to transform the double mutant, though these plasmids efficiently transformed cells harboring a hupA or hupB single mutation. The composite plasmid pFHS1, which consists of the f5 DNA fragment of F plasmid and the whole DNA of a pSC101 derivative that carries a temperature-sensitive mutation for DNA replication, was not stably maintained in the hup double mutant at 42°C. These findings strongly suggest that HU protein is required for ori2-dependent replication of the F plasmid.     

Distribution of Clostridium botulinum in Japan and in Shinkiang district of China

Soil specimens obtained from several areas of Japan, which are closely located to or facing the Continental land of China, were examined for the distribution of Clostridium botulinum, especially pertaining to types A and B. A total of 266 specimens of Japan, when cultured, showed no type A or B toxicity, although 30 (11.3%), 4 (1.5%), and 10 (3.8%) of the specimens showed C1, C2, and type E toxicities, respectively. On the contrary, types A and/or B toxicities were shown, by the same method, in 14 of 20 specimens of Shinkiang district, China. The highest number of C. botulinum cells found in one gram of soil specimen was 25 for type A and 10 for type B.     

Selective inhibition of high- but not low-affinity interleukin 2 binding by lectins and anti-interleukin 2 receptor alpha antibody

The present study demonstrated that various reagents can specifically reduce the affinity of high-affinity interleukin 2 receptor (IL-2R) but not that of low-affinity IL-2R. They included lectins such as wheat germ agglutinin (WGA), concanavalin A and phytohemagglutinin, and a chemical cross-linker, glutaraldehyde, in addition to anti-IL-2R monoclonal antibodies, HIEI and H-47. The affinity of the high-affinity IL-2R was reduced when the cells were treated with WGA or H-47 before, but not after, addition of 125I-labeled interleukin 2 (IL-2). Their inhibitory effects were also demonstrated by the chemical cross-linking method. On treatment with the reagents, the IL-2 binding to both IL-2R alpha and beta chains was inhibited and these inhibitory effects were seen only when the reagents were added before IL-2 addition, like their high-affinity reducing effects. These results support a supposition that the high affinity IL-2R is generated by assembly of the alpha and beta chains, and suggest that the IL-2 binding to IL-2R alpha and beta chains could induce stable constitution of the high-affinity state of IL-2R, but these affinity modulating reagents could perturb the optimum association between alpha and beta chains to generate the high-affinity IL-2R.