The ClpE protein involved in biogenesis of the CS31A capsule-like antigen is a member of a periplasmic chaperone family in Gram-negative bacteria

Abstract The putative chaperone-like protein ClpE, required for biogenesis of the Escherichia coli capsule-like antigen CS31A, was compared with ten known periplasmic chaperones from E. coli, Klebsiella pneumoniae, Bordetella pertussis, Haemophilus influenzae and Yersinia pestis . The amino acid sequence alignment was superimposed onto the three-dimensional structure of the PapD chaperone of uropathogenic E. coli , and amino acid residues involved in maintaining the structure integrity of the suggested binding site were found identical in most of the 11 chaperones. Construction of a phylogenetic tree to investigate the relationship within the chaperone family has revealed interesting degrees of relatedness between the different proteins.     

AICAR is not an endogenous mutagen in Escherichia coli

A number of observations in the Escherichia coli and Salmonella typhimurium literature could be explained by the hypothesis that a particular purine ribonucleotide precursor can be converted to the corresponding deoxyribonucleotide triphosphate, thereby becoming a base-analogue mutagen. The metabolite in question, AICAR (5-amino-4-carboxamide imidazole riboside 5′-phosphate), is also a by-product of histidine biosynthesis, and its (ribo)triphosphate derivative, ZTP, has been detected in E. coli. We constructed E. coli tester strains that had either a normal AICAR pool (pur ⁺ his ⁺ strains cultivated without purines or histidine) or no AICAR pool (purF hisG mutant strains, lacking the first enzyme of each pathway and cultivated in the presence of adenine and histidine). Using a set of lacZ mutations, each of which can revert to Lac⁺ only by a specific substitution mutation, we found that no base substitution event occurs at a higher frequency in the presence of an AICAR pool. We conclude that the normal AICAR pool in E. coli is not a significant source of spontaneous base substitution mutagenesis.     

A rapid assay for chloroplast-encoded triazine resistance in higher plants

We have designed a simple and rapid assay for chloroplast-based triazine resistance in higher plants using PCR amplification of thepsbA gene coupled toMaeI digestion of the amplified product to distinguish triazine resistant from sensitive biotypes. Our assay is universal and avoids the need of lengthy procedures of previously published assays, which either required spraying of seedlings in a controlled environment, quantification of chlorophyll fluorescence of leaf discs after incubation in triazine solution, DNA sequencing of thepsbA gene, or Southern-blot analysis. Our diagnostic system is qualitative, reliable, fast and simple. More than 100 seedlings taken directly from the field can be analyzed in one day. This system has a direct application towards a more rational use of herbicides in production fields. It also represents a valuable tool to monitor spreading of resistant biotypes through time and space and can serve as a model system applicable to other gene monitoring needs.     

Growth kinetics of Rhizopus arrhizus in solid state fermentation of treated cassava

Summary Growth kinetics of Rhizopus arrhizus MUCL 28168 were determined for different treatments of cassava during solid state fermentation. The best case gave a specific growth rate () of 0.24 h^-1, a yield calculated on a basis that oxygen consumption (Y_x/o) was 2.9 g biomass. g^-1 O₂ consumed and the maintenance coefficient (m) was 0.004 g O₂ consumed. g^-1 biomass. h^-1.     

EFFECTS OF LONG TERM EXPOSURE TO LOW TEMPERATURE ON THE PHOTOSYNTHETIC APPARATUS OF DUNALIELLA TERTIOLECTA (CHLOROPHYCEAE)1

The effects of long term exposure to suboptimal growth temperature on the photosynthetic apparatus of Dunaliella tertiolecta Butcher were investigated using carbon fixation rate versus irradiance curves and the variable fluorescence induction method. Carbon fixation rates per unite chlorophyll a at saturating (p^B_m) and subsaturating (α^B) irradiances were 55% and 39% lower, respectively, at 12° C than at 20° C. Chlorophyll a quotas and the spectrally averaged in vivo absorption cross section normalized to chlorophyll a (a^*) were not significantly different at these two temperatures. Analysis of the fluorescence kinetics revealed 1) no significant variations of the amount of PSII photoactive reaction centers per unit chlorophyll a, 2) a 14% decrease of the PSII quantum yield(+) and 3) a 29% decrease of the energy transfer efficiency between the light harvesting chlorophyll a pigment bed and the PSII reaction centers. The decrease in energy transfer efficiency between the antennae and the PSII reaction centers at 12° C was interpreted as a mechanism to avoid photoinhibition.     

Poly(amidoamine) dendrimer peripherally modified with 4-N,N-dimethylaminoethyleneamino-1,8-naphthalimide as a sensor of metal cations and protons.

A new poly(amidoamine) dendrimer from second generation whose periphery comprises sixteen fluorescent 4-N,N-dimethylaminoethylamino-1,8-naphthalimide units has been synthesized and characterized. In DMF, the dendrimer shows sensitivity to the presence of Cu(2+), Fe(3+) and protons. The changes in the fluorescence intensity of the material are in opposite directions if acids or metals are present. Fluorescence enhancements (FE from 5 to 9 depending on solvent) are recorded when the photoinduced electron transfer (PET) originating from the donating amine to the electron accepting naphthalimide is inhibited by the protonation of the N,N-dimethylamino groups. In the case of Cu(2+) cations, a fluorescence quenching (FQ of 6) is first observed, followed by fluorescence partial restoration. In the Fe(3+) case, the same behaviour is observed with a final FE of 2. The successive complexations of these cations by the dendrimer core and by the external rim of the dendrimer may explain the results.     

Freshwater Bacteria Can Methylate Selenium through the Thiopurine Methyltransferase Pathway

Involvement of the bacterial thiopurine methyltransferase (bTPMT) in natural selenium methylation by freshwater was investigated. A freshwater environment that had no known selenium contamination but exhibited reproducible emission of dimethyl selenide (DMSe) or dimethyl diselenide (DMDSe) when it was supplemented with an organic form of selenium [(methyl)selenocysteine] or an inorganic form of selenium (sodium selenite) was used. The distribution of the bTPMT gene (tpm) in the microflora was studied. Freshwater bacteria growing on 10 μM sodium selenite and 10 μM sodium selenate were isolated, and 4.5 and 10% of the strains, respectively, were shown by colony blot hybridization to hybridize with a Pseudomonas syringae tpm DNA probe. Ribotyping showed that these strains are closely related. The complete rrs sequence of one of the strains, designated Hsa.28, was obtained and analyzed. Its closest phyletic neighbor was found to be the Pseudomonas anguilliseptica rrs sequence. The Hsa.28 strain grown with sodium selenite or (methyl)selenocysteine produced significant amounts of DMSe and DMDSe. The Hsa.28 tpm gene was isolated by genomic DNA library screening and sequencing. BLASTP comparisons of the deduced Hsa.28 bTPMT sequence with P. syringae, Pseudomonas aeruginosa, Vibrio cholerae, rat, and human thiopurine methyltransferase sequences revealed that the levels of similarity were 52 to 71%. PCR-generated Escherichia coli subclones containing the Hsa.28 tpm open reading frame were constructed. E. coli cells harboring the constructs and grown with sodium selenite or (methyl)selenocysteine produced significant levels of DMSe and DMDSe, confirming that the gene plays a role in selenium methylation. The effect of strain Hsa.28 population levels on freshwater DMSe and DMDSe emission was investigated. An increase in the size of the Hsa.28 population was found to enhance significantly the emission of methyl selenides by freshwater samples supplemented with sodium selenite or (methyl)selenocysteine. These data suggest that bTPMT can play a role in natural freshwater selenium methylation processes.     

Evolution of reproductive systems in the genus Silene

The genus Silene contains both hermaphrodite, gynodioecious and dioecious species, dioecy being represented in three sections of the genus. To locate the events of change of reproductive systems, we compared ITS sequences of 22 species of Silene chosen throughout the whole genus, and four putative outgroup species. Gynodioecy, which is the most common reproductive system within the genus Silene and in closely related genera such as Saponaria and Dianthus, is proposed to be ancestral in the genus. Dioecy has evolved at least twice: once in the section containing S. latifolia, and once in the clade containing S. otites and S. acaulis ssp. bryoides. Evolution towards hermaphroditism, associated with evolution of selfing has also occurred at least twice, in S. gallica and S. comica.     

Classification and expression of a family of cyclin gene homologues in Brassica napus

In order to investigate the role of cell division in plant development, we isolated several plant genes which encode homologues of animal and yeast cell cycle regulators known as cyclins.Through the use of degenerate primers and the polymerase chain reaction (PCR) we isolated a Brassica sequence which showed homology to the cyclin box functional domain found within cyclin proteins. Southern blot analysis indicated that Brassica napus has a large number of genes containing cyclin box-related sequences. This was further supported by the isolation of cyclin box sequences from six different genomic clones. In addition, we have isolated two different cyclin cDNA clones, BnCYC1 and BnCYC2, from a Brassica napus shoot apical cDNA library. Both of the cDNA clones contain a destruction box regulatory domain similar to animal mitotic cyclins.Northern blot analysis using BnCYC2 shows mRNA levels which correlate well with the level of cell division in various tissues. Messenger RNA abundance was highest in 1–3 mm leaves, root tips and shoot apices. The mRNA detected using BnCYC1 was restricted to young leaves and the shoot apex, suggesting divergent, organ-specific roles for cyclin family members. The results demonstrate that the plant cyclin gene family is more extensive than previously demonstrated and consists of genes expressed in all dividing tissues as well as a subset of developmentally specific members.