Kinetics of alpha phosphate turnover in the free nucleotides of isolated rabbit hearts during the incorporation of adenosine or uridine]

The isolated rabbit heart was perfused with tritium-labelled adenosine or uridine at constant concentration (respectively 10 and 2 micrometer) and specific activity. The transfer rates of the precursors into free nucleotides were obtained by calculating the uptake of isotope into the products. The rates of phosphorylation of the nucleotides were estimated by following the incorporation of 32PO4 3- into the alpha phosphate groups. The results obtained with the two isotopes show that the rate of labelling of the alpha phosphate groups is a good estimate of the turnover of nucleotides when the mechanism of synthesis involves phosphorylation of a riboside.     

The saccus vasculosus of Anguilla anguilla (L.) from larva to adult

Summary The ultrastructure of coronet cells of the saccus vasculosus has been studied in specimens of Anguilla anguilla (L.) at different stages of its life cycle. At all the stages observed coronet cells are composed of a basal and an apical part, the latter bearing globules with primary vesicles. In the larva (a marine form) and in the fully metamorphosed small eel at the time of entry into freshwater the narrow lumen and the vesicles within the apical globules are filled with electron-dense material. In forms in which adaptation to freshwater has occurred, the saccus lumen appears expanded, the apical globules are better developed, and the electron-dense material has disappeared. It is suggested that the two situations observed represent different functional states of the organ, in relation to different conditions of environmental salinity.The authors gratefully acknowledge the contribution of Dr. G. Andreoli, of the University of Parma, who provided the Atlantic larvae for this study.     

A rapid method for determination of ribonucleotide reductase activity

Ribonucleoside diphosphate reductase activity is determined in centrifuged homogenates by following the conversion of cytosine ribonucleotide to cytosine deoxyribonucleotide. The enzymatic reaction is measured by monitoring the radioactivity of the reaction products separated by thin layer chromatography on PEI-cellulose plates. The method is rapid and permits the simultaneous processing of multiple samples.     

Paraoxonase 1 activity in the sperm-rich portion of boar ejaculates is positively associated with sperm quality

Associations of the activity of the paraoxonase 1 (PON1) enzyme with boar sperm quality still needs to be characterized, since boar ejaculates present distinct portions with differences in sperm concentration and quality. This study evaluated PON1 activity in the serum, in the distinct portions of boar ejaculates and estimated correlations with sperm quality parameters. Ejaculates and blood samples were collected from six boars for three weeks (two per week per boar; n = 36). Serum and post-spermatic portion PON1 activities were positively correlated (P = 0.01) but were both uncorrelated with the PON1 activity in the sperm-rich portion and in the whole ejaculate (P > 0.05). Differences in PON1 activity among boars were only observed in the sperm-rich portion of the ejaculate (P < 0.05). The PON1 activity in the serum and in the post-spermatic portion was generally negatively correlated with parameters of spermatozoa kinetics (P < 0.05). In the sperm-rich portion, PON1 activity was positively correlated with sperm concentration (P < 0.0001), curvilinear distance and velocity (both P < 0.05) and DNA integrity (P < 0.05), but negatively correlated with straightness and linearity (P < 0.05). Thus, boar ejaculates with increased PON1 activity in the sperm-rich portion may present increased concentration and spermatozoa with acceptable curvilinear velocity and distance and DNA integrity, which suggests that PON1 activity may be a biomarker for potential fertility.     

Gamma interferon is able to enhance the oxidative metabolism of human neutrophils

The oxidative metabolism of human neutrophils has been studied after incubation of the cells with recombinant interferon-y. Neutrophils incubated for 2-4 hours with 2-50 U/ml recombinant interferon-y undergo a higher respiratory burst measured both as Oz consumption and Oz- production when stimulated with formyl-methionyl-leucyl-phenylalanine, Concanavalin A or phorbol myristate acetate. The potentiating effect of interferon-y requires more than one hour of incubation, is optimal at 20-50 U/ml and depends on the presence of serum in the incubation medium. The interferon effect depends on new protein synthesis. Cycloheximide at doses which do not alter the respiratory response of normal cells completely prevents the potentiating effect of interferon.     

DNA ligase I is recruited to sites of DNA replication by an interaction with proliferating cell nuclear antigen: identification of a common targeting mechanism for the assembly of replication factories.

In mammalian cells, DNA replication occurs at discrete nuclear sites termed replication factories. Here we demonstrate that DNA ligase I and the large subunit of replication factor C (RF-C p140) have a homologous sequence of approximately 20 amino acids at their N-termini that functions as a replication factory targeting sequence (RFTS). This motif consists of two boxes: box 1 contains the sequence IxxFF whereas box 2 is rich in positively charged residues. N-terminal fragments of DNA ligase I and the RF-C large subunit that contain the RFTS both interact with proliferating cell nuclear antigen (PCNA) in vitro. Moreover, the RFTS of DNA ligase I and of the RF-C large subunit is necessary and sufficient for the interaction with PCNA. Both subnuclear targeting and PCNA binding by the DNA ligase I RFTS are abolished by replacement of the adjacent phenylalanine residues within box 1. Since sequences similar to the RFTS/PCNA-binding motif have been identified in other DNA replication enzymes and in p21(CIP1/WAF1), we propose that, in addition to functioning as a DNA polymerase processivity factor, PCNA plays a central role in the recruitment and stable association of DNA replication proteins at replication factories.     

Enhanced production of L-(+)-lactic acid in chemostat by Lactobacillus casei DSM 20011 using ion-exchange resins and cross-flow filtration in a fully automated pilot plant controlled via NIR

Due to the lack of suitable in-process sensors, on-line monitoring of fermentation processes is restricted almost exclusively to the measurement of physical parameters only indirectly related to key process variables, i.e., substrate, product, and biomass concentration. This obstacle can be overcome by near infrared (NIR) spectroscopy, which allows not only real-time process monitoring, but also automated process control, provided that NIR-generated information is fed to a suitable computerized bioreactor control system. Once the relevant calibrations have been obtained, substrate, biomass and product concentration can be evaluated on-line and used by the bioreactor control system to manage the fermentation. In this work, an NIR-based control system allowed the full automation of a small-scale pilot plant for lactic acid production and provided an excellent tool for process optimization. The growth-inhibiting effect of lactic acid present in the culture broth is enhanced when the growth-limiting substrate, glucose, is also present at relatively high concentrations. Both combined factors can result in a severe reduction of the performance of the lactate production process. A dedicated software enabling on-line NIR data acquisition and reduction, and automated process management through feed addition, culture removal and/or product recovery by microfiltration was developed in order to allow the implementation of continuous fermentation processes with recycling of culture medium and cell recycling. Both operation modes were tested at different dilution rates and the respective cultivation parameters observed were compared with those obtained in a conventional continuous fermentation. Steady states were obtained in both modes with high performance on lactate production. The highest lactate volumetric productivity, 138 g L(-1) h(-1), was obtained in continuous fermentation with cell recycling.