Enhanced production of pigment-free pullulan by a morphogenetically arrested Aureobasidium pullulans (ATCC 42023) in a two-stage fermentation with shift from soy bean oil to sucrose

A two-stage fermentation process was established for the production of pigment-free pullulan by the yeast-like fungus Aureobasidium pullulans (ATCC 42023). In the first stage, starting at pH 4.5 with soy bean oil as the carbon source and glutamate as the nitrogen source, a cell mass of about 15 g l^–1 dry cell weight was obtained, the population being restricted mainly to the yeast form of the microorganism (yeast form more than 90% of total cells) and the formation of pigment in the culture being prevented. Small amounts of pullulan (less than 2 g l^–1) are produced at this phase, and the viscosity remained low throughout the entire growth stage. When the oil and glutamate source were nearly exhausted (below 5% of initial amounts), the cells were shifted to a production stage with sucrose as the carbon source with continued nitrogen depletion. Production of pullulan started immediately with no lag period. During 50 h of the production phase more than 35 g l^–1 of pullulan was produced (productivity approx. 0.7 g l^–1), resulting in a large increase in the viscosity of the broth. The production yield of pollulan on the sugar was about 0.6 g g^–1. Morphogenesis from the yeast form of the microorganism to chlamydospores was still restrained and no pigment was formed in the culture during the production stage. A pigment-free polysaccharide, with a molecular mass in the range of 600–750 kDa, was recovered from the supernatant of the broth after solvent precipitation.     

Fluctuations of grass pollen content in the atmosphere of East Perugia and meteorological correlations (year 1989)

Summary Gramineae pollination from a pollen monitoring station located in the eastern suburb of Perugia and meteorological correlations are reported. The data refers to the year 1989. Grass pollen peak pollination was from May to July; in this period the influence of relative humidity and of temperature on pollen concentration was very high. Phenological observations, to identify the time of maximum stamen extension in the most common genera in the area, are also reported. During the period of investigation the counts of pollen grains over four-hour periods showed a regular diurnal rhythm with peaks of concentration in the four-hour period 16.00–20.00. Aerosporological data and meteorological data related to four-hour periods were correlated following different criteria.     

Peptide hormone-membrane interactions: the aggregational and conformational state of lipo-gastrin derivatives and their receptor binding affinity.

The (2RS)-1,2-dipalmitoyl-3-mercaptoglycerol/-, (2RS)-1,2-dimyristoyl-3-mercaptoglycerol/-, and (2RS)-1-myristoyl-2-palmitoyl-3-mercaptoglycerol/maleoyl-bet a-alanyl- [Nle15]-human-gastrin-(2-17) adducts were prepared as lipo-gastrin derivatives of explicitly primary amphiphilic properties. As representative of this class of lipo-gastrins, the dimyristoyl derivative has been thoroughly characterized in its aggregational state since, among the three compounds, theoretically it should exhibit the lowest degree of lipid character. It aggregates in aqueous solution to form monodispersed unilamellar spherical vesicles with dislocation of the peptide moiety at the bilayer surface in predominantly unordered structure. The liposomes are remarkably stable toward solubilization with trifluoroethanol and toward vesicle to micelle transition with neutral and negatively charged surfactants even above their critical micellar concentrations. Asymmetric fusion with the detergent micelles induces polydispersion of the liposomes in terms of shape and size without affecting in significant manner the mode of display of the gastrin portions at the bilayer surface. Only the positively charged hexadecyltrimethylammonium hydroxide provokes the collapse of the vesicles into mixed micelles with concomitant altered dislocation of the gastrin-peptide in the new aggregational state. Despite the lipid properties of the gastrin derivatives, i.e., formation of liposomes, they retain remarkable receptor affinities (IC50 = 1.5 x 10(-9) M for myristoyl-palmitoyl-gastrin, IC50 = 2.0 x 10(-9) M for di-myristoyl-gastrin and IC50 = 3.1 x 10(-9) M for di-palmitoyl-gastrin vs IC50 = 2.8 x 10(-10) M for Nle15-gastrin). Since the displacement of radiolabeled Nle15-gastrin from rat pancreatic acinar cell line membrane preparations by both the parent gastrin hormone and the three lipo-gastrins occurs in parallel manner, the data support a mechanism of receptor occupancy via accumulation of the gastrins at the membrane surface and their two-dimensional diffusion to the target receptor. Thereby the differentiated decrease of affinity in function of fatty acid chain length has to be attributed to the energetically more or less favored transfer of the monomers from the donor vesicles to the acceptor membranes. Moreover, according to this model migration of the lipo-gastrins with their interdigitating di-fatty-acyl moieties should be delayed, again in lipid structure-dependent manner, in comparison to the parent gastrin molecule, which is free to float in the membrane interfacial phase.     

Cytosine methylation in a CpG sequence leads to enhanced reactivity with Benzo[a]pyrene diol epoxide that correlates with a conformational change.

Benzo[a]pyrene (B[a]P) is a widespread environmental carcinogen that must be activated by cellular metabolism to a diol epoxide form (BPDE) before it reacts with DNA. It has recently been shown that BPDE preferentially modifies the guanine in methylated 5'-CpG-3' sequences in the human p53 gene, providing one explanation for why these sites are mutational hot spots. Using purified duplex oligonucleotides containing identical methylated and unmethylated CpG sequences, we show here that BPDE preferentially modified the guanine in hemimethylated or fully methylated CpG sequences, producing between 3- and 8-fold more modification at this site. Analysis of this reaction using shorter duplex oligonucleotides indicated that it was the level of the (+)-trans isomer that was specifically increased. To determine if there were conformational differences between the methylated and unmethylated B[a]P-modified DNA sequences that may be responsible for this enhanced reactivity, a native polyacrylamide gel electrophoresis analysis was carried out using DNA containing isomerically pure B[a]P-DNA adducts. These experiments showed that each adduct resulted in an altered gel mobility in duplex DNA but that only the presence of a (+)-trans isomer and a methylated C 5' to the adduct resulted in a significant gel mobility shift compared with the unmethylated case.     

Cross-species gene-family fluctuations reveal the dynamics of horizontal transfers

Prokaryotes vary their protein repertoire mainly through horizontal transfer and gene loss. To elucidate the links between these processes and the cross-species gene-family statistics, we perform a large-scale data analysis of the cross-species variability of gene-family abundance (the number of members of the family found on a given genome). We find that abundance fluctuations are related to the rate of horizontal transfers. This is rationalized by a minimal theoretical model, which predicts this link. The families that are not captured by the model show abundance profiles that are markedly peaked around a mean value, possibly because of specific abundance selection. Based on these results, we define an abundance variability index that captures a family''s evolutionary behavior (and thus some of its relevant functional properties) purely based on its cross-species abundance fluctuations. Analysis and model, combined, show a quantitative link between cross-species family abundance statistics and horizontal transfer dynamics, which can be used to analyze genome ‘flux’. Groups of families with different values of the abundance variability index correspond to genome sub-parts having different plasticity in terms of the level of horizontal exchange allowed by natural selection.     

D8S7 is consistently deleted in inverted duplications of the short arm of chromosome 8 (inv dup 8p)

Ten patients with inverted duplication of 8p (inv dup 8p) were studied with cytogenetic, biochemical and molecular techniques. The duplication for the region 8p12-p22 was always associated with a deletion of the locus D8S7 (mapped in 8p23.1) as demonstrated with the probe pSW50 by both in situ hybridization and Southern blot. Restriction fragment length polymorphisms detected by probes pSW50 (1 case) and by pG2LPL35 (locus LPL) (two cases) were informative as to a maternal origin of the anomaly. The activity of glutathione reductase, whose gene maps in the duplicated region at 8p21.1, was increased in all patients. The recognizable phenotype of inv dup 8p includes neonatal hypotonia, prominent forehead, large mouth with everted lower lip, abnormally shaped large ears, brain malformations and severe mental retardation. Our findings indicate that the chromosome rearrangement is homogeneous at least for the presence of the deletion and support the hypothesis of a common mechanism of origin.