Secretion of a 107 K dalton polypeptide into the medium from a haemolytic E. coli K12 strain

Summary Certain E. coli K12 strains are able to secrete a plasmid encoded 107 K protein into the culture medium. During exponential growth of the cells this protein represents approximately 1% of total cell protein.The presence of the 107 K polypeptide was demonstrated through the fortuitous use of strain MC4100. This gave a largely protein-free culture supernatant, presumably due to minimal lysis of whole cells. Pulse-labelling experiments showed that the secretion of the 107 K polypeptide reached a maximum during the stationary phase of growth, where it represented substantially more than 1% of total cell protein. The 107 K polypeptide is coded by the haemolytic plasmid pHly167, and appears to be related to a previously reported intracellular precursor form of the -haemolysin (Goebel and Hedgpeth 1982). However, additional extracellular factors appear to be required for -haemolysin activity since several nonhaemolytic mutants still secrete this protein.     

Isolation of streptothricin resistant mutants from E coli K12, strain A19

Mutants with various levels of resistance to streptothricin were isolated from Escherichia coli K12, strain A19 after mutagenesis with N-methyl-N-nitro-N-nitroso-guanidine and ethylmethane-sulfonate. Nourseothricin, a mixture of streptothricin F and D was the selection agent. Spontaneous resistant mutants could not be found. The streptothricin-resistant mutant E. coli A19 Stcr 2/2/1 shows cross-resistance to some of the aminoglycoside antibiotics investigated, but no cross-resistance to chloramphenicol and chlortetracyclin. These results indicate similar mechanisms of action of streptothricin and aminoglycoside antibiotics.     

Over-synthesis and instability of sigma protein in a merodiploid strain of Escherichia coli.

Summary We have used two different methods to study the rates of RNA polymerase subunit synthesis in haploid Escherichia coli K12, and a KLF10 rpoB,C ⁺ merodiploid derivative, when grown in glucose-minimal medium at 37°C. Our results indicate that the haploid strain produces , , and in the molar ratios, 1.01:0.99:2.90:0.26; and that all these subunits are reasonably stable during subsequent growth. The merodiploid produces at the same rate as the haploid, and at a 42% higher rate, and sigma at twice the rate. Some 40% of the newly synthesised and is degraded within one hour; the residuum is as stable as in the haploid. is stable throughout. By contrast, sigma is subject to a marked and continuous turnover in the merodiploid. These results are discussed in terms of gene dosage and regulatory effects.     

Cell division and DNA synthesis in uvrA recA double mutants of E. coli K12

Summary Cell division and incorporation of ³H-thymidine into acid-insoluble fraction were investigated for three uvrA recA double mutants of E. coli K12 irradiated with UV at 1.5 ergs/mm², producing about ten pyrimidine dimers per genome (about 0.01% survival). Cell division was measured both in M9 medium and in the same medium which was made very viscous by the addition of Metlose (the same product as Methocel used by Lin et al., 1971). It was found that a major fraction of irradiated bacteria continues to divide once or twice and stops thereafter. Incorporation of ³H-thymidine proceeded at a considerable rate for a short period following irradiation and then stopped. During subsequent incubation, the incorporation gradually decreased and after 4 h incubation most of the early incorporated radioactivity disappeared from the acid-insoluble fraction. These results indicate that cell division occurs after irradiation without parallel DNA synthesis as in a recA thy mutant of E. coli K12 deprived of thymine (Inouye, 1971). These results suggest that UV irradiation increases lethal sectoring due to the reckless cell division without parallel DNA synthesis. Since DNA synthesis took place only for a short period after irradiation, it may be assumed that the recA gene normally has at least a dual function; 1. elimination of damage induced by UV to support elongation or initiation of DNA, and 2. maintenance of coordination between DNA synthesis and cell division.     

Synthesis of paf-acether by E. coli K12

Paf-acether (platelet-activating factor) is one of the most potent mediator of inflammation released from and acting on most cells that participate in inflammatory diseases. Its molecular structure is 1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholine. Two metabolic steps are involved in its biosynthesis: the action of a phospholipase A2 on choline-containing membrane alkyl-ether lipids results in the production of lyso paf-acether and acetylation of the lyso compound by an acetyltransferase yields the biologically active molecule. Membrane alkyl-ether lipids can therefore be considered as potential precursors of paf-acether and their composition has been studied in various cell types. In this work, we investigated the presence of paf-acether in E. coli. Our results showed that paf-acether can be obtained from E. coli K12 under a variety of bacterial growth conditions. Paf-acether from E. coli exhibited the same physicochemical and biological characteristics as synthetic paf-acether and that from eucaryotic cells. Therefore, it appears that E. coli itself has the ability of producing paf-acether, a result that could be of some importance with respect to the pathogenesis of Enterobacteria and the use of E. coli in the recombinant DNA technology.     

Ultrastructure of LLR-mutants of E. coli K12]

The author studied the ultrastructure of two spherical E. coli K12 mutants (llr) obtained under the effect of N-nitroso-N-methylurea. Seven morphological types of cells differing from one another by shape, size and cytoarchitectonics were distinguished. Superficial structures of the majority of the cells were represented by the membranes of the cell wall and the cytoplasmic membrane of common structure. Some of the cells had only one membrane coat and a high electron optic density of the cytoplasm. Transitional forms of cells were also encountered. The ultrastructure of each morphological type in the population of the llr-mutants was described in detail. The capacity of the mutants to vacuolization, to the intra- and extracellular budding, and also the ability to form multiple membrane structures resembled analogous structures of stable L-forms of the Gram-negative microbes. The problems of morphological differentiation of the L-forms and of the llr-mutants, and also problems connected with the formation of the multiple membrane structures and small elemental bodies in the cells of the llr-mutants are discussed.     

DNA synthesis in F- cells treated with filtrates of male strains of E. coli K 12

Summary Filtrates of male strains of E. coli K12 partially inhibit DNA synthesis in F ^– cells and do not influence RNA and protein syntheses.     

Characterization of the E.coli K12 strain AB1157 as impaired in guanine/xanthine metabolism

The widely used E. coli K12 strain AB1157 is impaired in guanine (xanthine) metabolism. Mutants blocked in purine biosynthesis before the stage of inosine monophosphate synthesis do not grow on external guanine or xanthine.The genetic nature of the Gua/Xan lesion is a deletion in the chromosome that covers the pro A gene. The lesion causes reduced uptake of guanine.