H. Braná

Splitting of the cyclic 3′, 5′-adenosine monophosphate in cell-free system ofEscherichia coli

Broken cells ofEscherichia coli contain an enzyme system breaking down cyclic 3′,5′-adenosine monophosphate (Ado-3′,5′-P). The enzyme splitting this nucleotide is located in the supernatant fraction at 20,000 ×g. Some characteristics of the enzyme were studied. In contrast with the animal enzyme theEscherichia coli enzyme is not inhibited by caffeine.

Adenyl cyclase in cell-free extracts ofEscherichia coli

The adenyl cyclase enzyme system was detected in the cells ofEscherichia coli disrupted by sonic treatment. This enzyme activity is located mainly in the cell fraction sedimenting at 2,000×g, i.e. in the cytoplasmic membrane fraction. A prolonged sonication treatment of the cell suspension was followed by the disappearance of activity in the membrane preparation. The pH optimum of the adenyl cyclase inEscherichia coli was on the alkaline side, around pH 9.

Stability of the hybrid plasmid pIM138 and its curing by some eliminating agents.

Hybrid plasmid pIM138 was constructed by insertion of a chromosomal fragment with the threonme operon fromEscherichia coli into the pBR322 vector. Molar mass of pIM138 was 2.8 Mg/mol. Heteroduplexes between pBR322 vector and pIM138 hybrid DNA molecules were prepared. The hybrid plasmid shows a high stability against the curing effect of rifampicin and clorobiocm inE. coli SK1590thr host.

The effect of actinomycin D and flavomycin onEscherichia coli R+ strains

When tested by14C-uracil incorporation, a higher permeation of actinomycin D into R+Escherichia coli cells was observed. From actinomycin D and flavomycin only flavomycin was found to be effective in R+ cells selective growth inhibition. The results indicate that the effect of flavomycin is related to the fertility functions of the strain. The possible practical importance of flavomycin application for R+ cells elimination in the bacterial population is discussed.

A hydrogen sulphide producing Gram-negative rod from water.

The biochemical properties of a hydrogen sulphide-producing Gram-negative rod, provisionally designated HG group, were compared with those of the known H2S-producing and H2S-negative Enterobacteriaceae and related organisms. Sixty-four tests were used as a basis for numerical identification. All these tests demonstrated a distinctness of the HG group from other members of Enterobacteriaceae and related organisms. Results of numerical identification are discussed. According to the guanine-plus-cytosine molar content in DNA the bacterium could belong to the tribe Escherichiae of the family Enterobacteriaceae. Plasmids of different molecular size or linear fragments of DNA were found in 17 of 19 strains which indicates that the H2S production is not in correlation with the occurrence of a plasmid of definite size. So far, the only habitat of the HG group had been water, and it seems to be no rarity. Among 28 HG strains a single isolate HG16 was found which differs from HG group in biochemical properties. The distinctness of this single isolate has been confirmed also by numerical identification. Note: On the basis of DNA-DNA hybridization performed by Dr. P.A.D. Grimont and coworkers the HG group has been established as a new genus and a single species. The authors accordingly propose for the group the generic name Budvicia and the specific epithet aquatica.

Physical characterization of the plasmid pON5300

SummaryThe antibiotic resistance plasmid Rldrd19Km-which has spontaneously lost its kanamycin resistance marker, and its derivative pON5300, were analysed using the restriction endonucleases SalI, BamHI, HindIII and EcoRI. The fragment patterns were compared with that of the Rldrd19 and the fragments responsible for kanamycin resistance were found to be missing in Rldrd19Km− and pON5300. In these plasmids a 7 Mg/mol EcoRI fragment was observed instead of the D (6.3 Mg/mol) fragment of Rldrd19. Further a new 6 Mg/mol EcoRI restriction fragment was observed in pON5300. Using double digestions it was shown that the new fragment does not carry restriction sites for HindIII, BamHI and SalI endonucleases. The non-homology of the analysed plasmid was proved electron microscopically by heteroduplex techniques. The possibility of amplification in the regulatory region for the expression of R-determinants in pON5300 is discussed.

Mutation affecting normal colony formation inEscherichia coli K-12

The effect of a mutation in a gene located near thepro marker and determining the normal colony formation and the cell density inEscherichia coli has been analyzed. Thencf mutation has no effect on the permeability of the cell to actinomycin D. On the other hand, a higher level of ATP and a defect in the accumulation of the reserve material in the mutant cell were detected.

A special deoxyribonuclease activity accompanying competence-inducing activity

A deoxyribonuclease activity accompanies the competence substance isolated from transformableDiplococcus pneumoniae even in well purified fractions. The deoxyribonuclease seems to exhibit a rather different kind of activity from the one found as a major nuclease in a partially purified competence substance. The products of interaction between the enzyme and double-stranded DNA would indicate that the enzyme might act as an “unwindase” on the double-stranded DNA.

The adenosine triphosphate pool and the adenyl cyclase activity inEscherichia coli strains with different UV sensitivity

Two genetically relatedEscherichia coli strains were tested for the ATP pool and for the adenyl cyclase activity in the cell membrane fractions. TheEscherichia coli strain BS1with a high UV sensitivity showed a higher ATP level than the wild typeEscherichia coli B strain. The adenyl cyclase activity was found to be lower in the sensitive strain than in the wild typeEscherichia coli.

The substance with competence inducing activity interacts with DNA and produces a hyperchromic increase of optical density.

The active substance inducing competence in non-competent cells ofDiplococcus pneumoniae was isolated by ammonium sulphate precipitation from the sterile filtrate of a culture 40–60 min after the peak of competence had passed. After being dialyzed and dried from the frozen state the substance was further purified by chromatography on Sephadex G100 or DEAE cellulose column. In all cases the fractions containing the competence inducing activity contained also the transforming DNA-inactivating activity. A hyperchromic increase of optical density was found on incubating the active substance with various DNAs and RNAs as substrates. The hyperchromic increment is different from that of pancreatic endo-nuclease I and phosphodiesterase.