Inhibitory effect of cortisone acetate on the stimulation of rat liver cytosol l-serine Pyruvate aminotransferase by dibutyryl adenosine 3′,5′-monophosphate

An injection of cortisone acetate at a dose of 5 mg/100 g body weight concomitant with dibutryl cyclic AMP prevents the increase in the activity of rat liver cytosol serine aminotransferase (L-serine: pyruvate aminotransferase, EC 2.6.1.51) elicited by the nucleotide with a lag of about 2 h. If the glucocorticoid is given 2 h prior to the nucleotide inducer, the lag disappears. The inhibitory effect of cortisone acetate gradually decays and is no longer detectable 12 h following its administration. Theophylline, insulin and glucose at doses which affect significantly the level of tyrosine aminotransferase, have no effect on the level of serine aminotransferase and on the cortisone inhibition. The inhibitory effect of the glucocorticoid on the dibutyryl cyclic AMP-mediated increase in serine aminotransferase diminishes with the age of animals. Increase in the enzyme activity by a single dose of glucagon can also be inhibited by cortisone acetate and actinomycin D as in the case with dibutyrl cyclic AMP as an inducer. The possibility of the existence of a specific inhibitory factor which is formed in response to cortisone acetate is discussed.     

Quadruplex and pentaplex self-assemblies of oligonucleotides containing short runs of 8-aza-7-deaza-2'-deoxyisoguanosine or 2'-deoxyisoguanosine.

Oligonucleotides containing 8-aza-7-deaza-2'-deoxyisoguanosine (4) were investigated regarding their self-assembly in aqueous solution. The aggregation of 4 was compared with that of oligonucleotides containing 2'-deoxyisoguanosine (2b) and 2'-deoxyguanosine (1b). For this purpose the phosphoramidite of 4 was synthesized which was protected by a dibutylaminomethylidene residue at the amino group and a diphenylcarbamoyl residue at the 2-oxo function. Solid-phase synthesis furnished oligonucleotide containing short runs of the nucleoside 4. The self-assembly of the oligonucleotide 5'-d(T(4)4(4)T2) was studied by ion-exchange chromatography. The formation of a pentaplex was observed in the presence of Cs+, while a tetraplex is formed when the counter ion is Na+ or Rb+. The cation selectivity of the oligonucleotide 5'-d(T(4)4(4)T2) was found to be different from the parent 5'-d(T(4)isoG(4)T2) which was forming the tetraplex as well as a pentaplex in aq RbCl solution.     

Construction of chlorobenzene-utilizing recombinants by progenitive manifestation of a rare event

Separate continuous cultures of Pseudomonas putida R5-3, grown on toluene, and Pseudomonas alcaligenes C-O, grown on benzoate, were concentrated and continuously amalgamated on a ceramic bead column, which was subjected to a continuous stream of chlorobenzene vapors. A recombinant strain, P. putida CB1-9, was isolated in less than 1 month. P. alcaligenes C-0 grew on benzoate and 3-chlorobenzoate but not on toluene, P. putida R5-3 grew on benzoate and toluene but not on 3-chlorobenzoate, and neither strain grew on chlorobenzene or 1,4-dichlorobenzene; however, the recombinant P. putida CB1-9 grew on all of these substrates. Chlorobenzene-utilizing strains were not found in continuous cultures run at the lowest growth rate (0.05/h) or in the absence of the donor strain, P. alcaligenes C-0. Chloride was released in stoichiometric amounts when P. putida CB1-9 was grown on either chlorobenzene or 1,4-dichlorobenzene. The recombinant strain was related to P. putida R5-3, phenotypically and genetically. Restriction enzyme digests of the single 57-kilobase (kb) plasmid in R5-3 and of the single 33-kb plasmid in CB1-9 were similar, but also indicated rearrangement of plasmid DNA. Coincidental or causal to the loss of the 24-kb fragment was the observation that the recombinant--unlike its parent, R5-3--did not grow on xylenes or methylbenzoates. Although both ortho-pyrocatechase (OP) and meta-pyrocatechase (MP) were found in CB1-9 and R5-3, MP activity was 20- to 50-fold higher in R5-3 cells grown on 4-methylbenzoate than in the same cells grown on benzene.(ABSTRACT TRUNCATED AT 250 WORDS)     

Microcalorimetric investigation into the metabolic activity of rat caecal flora in the presence of different sugars and sugar substitutes

When adapting young rats to different sugar substitutes (sorbitol, PolydextroseR and PalatinitR), effects were seen in the caecal morphology and caecal content e.g. bacterial concentration, which did not occur when adapting rats to sugars (glucose, sucrose). For in vitro studies, anaerobic growth of caecal flora in thioglycollate medium with and without the respective substances was monitored by continuous measurement of heat production, optical density and pH. Additionally, biochemical analyses and light microscopic observations were performed in order to detect differences between adapted and non-adapted flora. In particular the microcalorimetric data furnished valuable information about alterations in bacterial metabolic activity after adaptation to sugars and sugar substitutes, and clearly indicated that all the substances tested influenced the metabolism of caecal flora.     

Separation of multiple components of HeLa cell nuclear extracts required for pre-messenger RNA splicing

Components essential for nuclear pre-messenger RNA splicing have been partially purified from HeLa cell nuclear extracts by chromatography on DEAE-Sepharose, heparin-Sepharose, Mono Q, and Mono S. We have obtained six fractions which, when combined, efficiently splice a synthetic adenovirus 2 major late RNA substrate in vitro. All fractions contain components that support the formation of splicing intermediates (the cleaved 5' exon and the intron-exon 2 lariat). At least one of the fractions also contains an activity that is essential for the second step in the splicing reaction, namely cleavage at the 3' splice site and exon ligation. Two of the fractions are enriched in the major small nuclear ribonucleoprotein particles U1, U2, U4/U6, and U5. They participate in the formation of the splicing complexes which precedes the cleavage and ligation reactions. The remaining four fractions appear to contain protein factors, as suggested by their resistance to micrococcal nuclease.     

Structural elements of bactopterin from Pseudomonas carboxydoflava carbon monoxide dehydrogenase

Bactopterin is a novel pterin occurring in bacterial molybdoenzymes as the organic portion of the molybdenum cofactor. Its structure is investigated here. The compound contains a single pterin ring and carries a side chain at carbon atom 6 of the pterin nucleus as indicated by the formation of pterin-6-carboxylic acid upon alkaline permanganate oxidation. Studies with phosphate-cleaving enzymes revealed the presence of two monophosphoric acid monoesters. The affinity of reduced bactopterin for thiol-Sepharose points to the presence of thiol(s) in active bactopterin.     

Characterization of the Ss sialoglycoprotein and its antigens in Rhnull erythrocytes

The Ss sialoglycoprotein (glycophorin B) and its antigens in Rhnull erythrocytes, which lack the Rhesus blood group antigens, due to apparently silent (amorphic type) or independent suppressor (regulator type) genes, were investigated. The quantity of the molecule in amorphic and in regulator type red cell membranes was found to be decreased by about 60%-70%, as judged from sodium-dodecylsulfate polyacrylamide gel electrophoresis. The Ss glycoprotein content in the erythrocytes from heterozygotes (regulator type) was diminished to an extent of about 30%. Confirming and extending previous studies, the S, s, Ux, Uz and 'N' antigens were slightly weakened in Rhnull erythrocytes. The U and Duclos receptors were only slightly or not depressed in amorphic Rhnull cells, but almost absent from or not detectable in those of the regulator type. This demonstrates that an additional alteration, apart from the decreased Ss glycoprotein content of the membranes, accounts for the weakness of these receptors in regulator type cells. We propose the hypothesis that (a) protein(s) encoded by the Rhesus locus form(s) a complex with the Ss glycoprotein. Thus, it (they) might facilitate the incorporation of the Ss glycoprotein into the membrane and also contribute to the complete expression of the U and Duclos antigens in normal cells.     

Testing the “methanochondrion concept”: are nucleotides transported across internal membranes in Methanobacterium thermoautotrophicum?

In order to test the Methanochondrion concept, uptake of adenine nucleotides in various membrane preparations of Methanobacterium thermoautotrophicum was studied. The uptake showed properties which are in general interpreted as indicative of a transport mechanism: (i) kinetics in the time range of minutes, (ii) temperature dependence, (iii) substrate specificity and (iv) failure to remove the substrate by extensive washing.However, nucleotide transport as an interpretation of this uptake can definitely be excluded. Not only an exchange mechanism of the mitochondrial type, but also a general exchange or an uniport mechanism was ruled out. In contrast, the nucleotide uptake was shown to be actually a tight and specific binding of ADP and ATP to binding sites at the interior side of the cell membrane. This was conclusively demonstrated in protoplasts obtained from M. thermoautotrophicum cells. In these protoplasts which do not contain internal membranes also nucleotide binding was observed, but only after disruption of the plasma membrane by osmotic lysis, which leads to the exposure of binding sites.