Stepwise cloning and genetic organization of the seemingly unclonable HgiCII restriction-modification system from Herpetosiphon giganteus strain Hpg9, using PCR technique.

The genes, hgiCIIR and hgiCIIM, that encode the HgiCII restriction and modification (R-M) system from Herpetosiphon giganteus strain Hpg9, an AvaII isoschizomer recognizing the sequence, GGATCC, were cloned in Escherichia coli. Cloning the respective hgiCIIM gene was achieved via in vitro selection both from a Sau3AI- and an NheI-generated plasmid gene library using AvaII, a commercially available isoschizomer of HgiCII. However, all attempts to clone the closely linked hgiCIIR and M genes in a single step resulted in deletions spanning parts of the coding region of hgiCIIR. Therefore, cloning of the missing 3'-terminal part of this gene was achieved by applying the inverse polymerase-chain-reaction technique. All attempts to construct an enzymatically active R.HgiCII failed; only the inactivated hgiCIIR gene could be cloned. Sequencing of the hgiCIIRM region (carrying predesigned small mutations in the R gene) disclosed three open reading frames (ORFs): one small ORF preceding the methltransferase (MTase)-encoding gene, plus those encoding M.HgiCII (49,620 Da) and R.HgiCII (30,891 Da). M.HgiCII exhibits the common motif of ten conserved amino-acid blocks typically found within the group of m5C-MTases. The R-M system of HgiCII reveals strong homologies to the isoschizomeric R-M system of HgiBI from H. giganteus strain Hpg5, which, in contrast, could be cloned in one step.     

A polysensory pathway to the forebrain of the pigeon: the ascending projections of the nucleus dorsolateralis posterior thalami (DLP).

Visual evoked potentials (VEPs) in the associative neostriatum caudolaterale (NCL) have shorter latencies than those recorded in other visual forebrain areas. Therefore visual input into NCL probably stems from a subtelencephalic relay. Tracing experiments revealed a projection of the nucleus dorsolateralis posterior thalami (DLP) into those portions of NCL in which visual, auditory, and somatosensory afferents from intratelencephalic parasensory areas terminate. Since VEPs in NCL are abolished after DLP-lesions, this structure has to be the critical relay. However, DLP also projects to other associative forebrain areas and parts of the basal ganglia. Previous experiments had furthermore revealed that DLP-neurons integrate visual, auditory, and somatosensory inputs. Thus, the DLP-projection onto various associative forebrain areas represents a true polysensory thalamotelencephalic system.     

Photosynthesis ofChlorella vulgaris as affected by diaqua(4-chloro-2-methylphenoxyacetato)copper(II) complex

Inhibitory effect of diaqua(4-chloro-2-methylphenoxyacetato)copper(II) complex (MCPACu) on oxygen evolution rate in algaeChlorella vulgaris was found. Electron paramagnetic resonance and fluorescence spectroscopy showed that the sites of action of MCPACu are Z⁺ and Y⁺ intermediates on the donor side of photosystem 2 as well as the manganese cluster in the oxygen evolving complex.     

Seasonal variations in the level of immunoglobulins and serum proteins of children differing by exposure to air-borne lead

A semicohort of children, initial age about 11.5 years, from an exposure area near a secondary lead smelting plant (E group children) was examined for some humoral immune response parameters in the blood and saliva and compared to a group of control children matched by age living in a relatively unpolluted rural area (Co group children). All examinations were performed every autumn and spring over a period of 2 successive years, and the data included in the final analysis were only from children who had completed at least 3 of the 4 examination series performed. To sum up, children from area E showed a clearcut elevation of mean blood lead (Pb) levels with marked seasonal fluctuations peaking in spring; Pb levels in the blood of Co group children had no such season-related fluctuation patterns and tended to slightly decline over the two years of observation. Delta-aminolevulinic acid dehydratases (ALAD) levels in area E were of distinctly lower mean values than in area Co, but no correlation with mean blood Pb levels could be observed. Levels of IgG were more or less inversely correlative with Pb levels in both sexes, levels of IgM showed a distinct downward trend in E group children at all samplings. Total serum protein (SP) levels of children from area E showed a reverse correlation with plumbemia, secretory IgA (sIgA) concentrations showed a decrease in E group children of both sexes, but the values tended to converge with control values. Levels of alpha 1-antitrypsin (A 1-AT) showed marked season-related fluctuation patterns in Co children, the curves in E group children turned steeply upward from the third examination series on. Ceruloplasmin (CPL) levels in E group children had a clearcut upward trend at three subsequent blood samplings, the curves in Co group children continued to drop. Children from area E showed markedly higher frequency rates of abnormal values than did children from Co area.     

Functional reconstitution of carrier proteins by removal of detergent with a hydrophobic ion exchange column

A method has been developed for the functional reconstitution of membrane proteins in phospholipid vesicles. This method is an extension of a previously published procedure (Ueno, M., Tanford, C. and Reynolds, A. (1984) Biochemistry 23, 3070-3076) for the formation of unilamellar vesicles from mixed micelles of egg phosphatidylcholine and dodecyl octaoxyethylene ether. Mixed micelles are formed from detergent-solubilized protein and egg-yolk phospholipid vesicles. These micelles are subjected to repeated passage through small columns filled with Amberlite XAD-2 beads. Several carrier proteins from the inner mitochondrial membrane have been reconstituted in this way; experimental data are shown for the aspartate/glutamate carrier and the ADP/ATP carrier. Certain parameters proved to be important for optimal efficiency of reconstitution: the ratio of detergent/phospholipid in the mixed micelles, the concentration of phospholipid during the hydrophobic chromatography, the ratio of phospholipid/protein, (d) the ratio of detergent/Amberlite XAD 2 beads, the number of column passages, and the type of detergent. After optimization of these parameters, phospholipid vesicles with a diameter of about 150 nm were obtained. The main advantage of this procedure, however, lies in the fact that high amounts of membrane protein can be incorporated into the phospholipid vesicles, i.e. up to 15% (w/w).     

Biology of DNA restriction.

Our understanding of the evolution of DNA restriction and modification systems, the control of the expression of the structural genes for the enzymes, and the importance of DNA restriction in the cellular economy has advanced by leaps and bounds in recent years. This review documents these advances for the three major classes of classical restriction and modification systems, describes the discovery of a new class of restriction systems that specifically cut DNA carrying the modification signature of foreign cells, and deals with the mechanisms developed by phages to avoid the restriction systems of their hosts.     

Behavior of hydroxyproline in the rat lung following exposure to surface dust and industrial ashes

There was analysed, by means of biochemical examinations, the behaviour of hydroxyproline level in the lungs of experimental animals exposed to the action of soil dusts and electroenergetic ashes. 44 female white rats of Wistar strain were used in the experiment. The animals were administered intratracheally 0.2 ml dusts and electroenergetic ashes suspension. After decapitation, prepared material was taken from the left lung. Hydroxyproline determinations were performed according to Prokop's und Undenfried's method modified by Tomaszewski and Hanzlik. 9 months after the administration of the dusts, the contents of hydroxyproline slightly differed from the levels observed in control group.     

Soil dust as a factor which changes the histochemical picture of the rat lung

White Wistar female rats only once were administered intratracheally a suspension of the soil dust in the physiologic saline. The respirable fraction of the soil dust was used for preparing the suspension of the latter. The experiment was carried out in 2 groups of rats. The animals of the 1st group received the soil dust suspension which was the mixture of the soil samples from 3 different places in Poland, the animals of the 2nd group from 3 other places in Poland. The control animals were once administered the physiologic saline. 9 months after the application of dust, the specimen of the left lung were taken for histochemical examinations. The reactions to the activity of succinic and lactate dehydrogenase, of tetrazole reductase NADP, and of acid phosphatase and ATP-ase were performed. The changes in the activity of the studied respiratory enzymes and the increase of the reaction to the acid phosphatase were observed. The results reveal the disorders of the oxidation-reduction processes and the intensification of the lytic processes in the pulmonary parenchyma of the animals exposed to the action of the soil dust.