The influence of temperature on glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase and the regulation of these enzymes in a mesophilic and a thermophilic cyanobacterium

The activities and kinetics of the enzymes G6PDH (glucose-6-phosphate dehydrogenase) and 6PGDH (6-phosphogluconate dehydrogenase) from the mesophilic cyanobacterium Synechococcus 6307 and the thermophilic cyanobacterium Synechococcus 6716 are studied in relation to temperature. In Synechococcus 6307 the apparent K _m's are for G6PDH: 80M (substrate) and 20M (NADP⁺); for 6PGDH: 90M (substrate) and 25M (NADP⁺). In Synechococcus 6716 the apparent K _m's are for G6PDH: 550M (substrate) and 30M (NADP⁺); for 6PGDH: 40M (substrate) and 10M (NADP⁺). None of the K _m's is influenced by the growth temperature and only the K _m's of G6PDH for G6P are influenced by the assay temperature in both organisms. The idea that, in general, thermophilic enzymes possess a lower affinity for their substrates and co-enzymes than mesophilic enzymes is challenged.Although ATP, ribulose-1,5-bisphosphate, NADPH and pH can all influence the activities of G6PDH and 6PGDH to a certain extent (without any difference between the mesophilic and the thermophilic strain), they cannot be responsible for the total deactivation of the enzyme activities observed in the light, thus blocking the pentose phosphate pathway.Abbreviations G6PDH glucose-6-phosphate, dehydrogenase - 6PGDH 6-phosphogluconate dehydrogenase - G6P glucose-6-phosphate - 6PG 6-phosphogluconate - RUDP ribulose-1,5-bisphosphate - Tricine N-Tris (hydroxymethyl)-methylglycine     

Regulation of the synthesis of lutropin-induced protein in rat testis Leydig cells

The mechanism of action of lutropin on the stimulation of the synthesis of a specific lutropin-induced protein in rat testis Leydig cells was investigated. Lutropin-induced protein has a mol.wt. of approx. 21000 and is detected by labelling the Leydig-cell proteins with [³⁵S]methionine, followed by separation by polyacrylamide-gel electrophoresis and radioautography of the dried gel. The incorporation of ³⁵S into lutropin-induced protein was used as an estimate for the synthesis of the protein. Incubation of Leydig cells with dibutyryl cyclic AMP or cholera toxin also resulted in the stimulation of synthesis of the protein. Synthesis of lutropin-induced protein, when maximally stimulated with 100ng of lutropin/ml, could not be stimulated further by addition of dibutyryl cyclic AMP. Addition of 3-isobutyl-1-methylxanthine, a phosphodiesterase inhibitor, further increased synthesis of the protein in the presence of a submaximal dose of lutropin (10ng/ml) but not in the absence of lutropin or with maximal amounts of lutropin (100 and 1000ng/ml). Actinomycin D prevented the effect of lutropin on the stimulation of lutropin-induced protein synthesis when added immediately or 1h after the start of the incubation, but not when added after 5–6h. This is interpreted as reflecting that, after induction of mRNA coding for lutropin-induced protein, lutropin had no influence on the synthesis of the protein in the presence of actinomycin D. Synthesis of the protein was also stimulated in vivo by injection of choriogonadotropin into rats 1 day after hypophysectomy, and the time course of this stimulation of lutropin-induced protein synthesis in vivo was similar to that obtained by incubating Leydig cells in vitro with lutropin. From these results it is concluded that stimulation of lutropin-induced protein synthesis by lutropin is most probably mediated by cyclic AMP and involves synthesis of mRNA.     

ACTH-induced inhibition of endogenous rat brain protein phosphorylation in vitro: Structure activity

ACTH_1–24 inhibits the endogenous phosphorylation in vitro of distinct SPM protein bands. Using N-terminal fragments of ACTH, the structure-activity requirements for this effect were studied. A rather complex interaction of the ACTH fragments with endogenous SPM phosphorylation was observed. The effects were not only dependent on the primary structure of the peptide used, but also on the protein band studied and the ATP/SPM ratio used in the incubation system. ACTH_1–24 did not interfere with the ATP-hydrolyzing activity of the SPM preparation, nor did it influence the endogenous phosphatase activity. Therefore, a direct interaction of ACTH with SPM protein kinase(s) is likely to be responsible for its effect on phosphorylation.     

The origin of mentha — gracilis (Lamiaceae). II. essential oils

Essential oils were eamined in nine clones of Mentha arvensis, four clones of M. spicata, and 20 clones of M. gracilis. An F₁ hybrid of M. arvensis M. spicata, selected on the basis of morphology and chromosome number, was matched with one clone of M. Gracilis. Genes for the inheritance of limonene, 1,8-cineole, linalool, isomenthone, carvone, and piperitenone oide were identified in one clone of M. arvensis and two clones of M. spicata. The range of essential oil compounds detected indicates that no one character can be used to identify M. gracilis, but the critical compounds of the oil of M. gracilis can be derived from crosses of M. arvensis M. spicata.     

Stress response induced by DNA damage leads to specific, delayed and untargeted mutations

Summary Cells of the mouse T-lymphoma line GRSL13 were treated with 8-methoxy-psoralen plus longwave ultraviolet light (PUVA) under conditions where the biological effects are mainly due to non-persistent DNA crosslinks (PUVA-CL treatment). Fluctuation analysis showed that PUVA-CL treatment resulted in an enhancement of the mutation rate in the progeny of treated cells, which persisted until the eleventh generation after treatment. Since only 5 cross-links are available to account for 52 mutational events observed in the coding region, about 90% of the induced mutational events must have been untargeted. This was confirmed by molecular analysis of these mutations, which showed that 53% of the point mutations arose at sites which are not a target for psoralens. This supports the hypothesis that stress responses may give rise to untargeted mutagenesis. Further support for this hypothesis is provided by the observation that 8-methoxy-psoralen (8-MOP) or UVA alone (both of which are known to induce many pleiotropic effects) each acted as indirect mutagen by enhancing the mutation rate 2–4 fold in the progeny of treated cells.     

Enzymatic preparation of 1,6-anhydro-muropeptides by immobilized murein hydrolases from Escherichia coli fused to staphylococcal protein A

Summary In order to produce biologically active 1,6-anhydro-muropeptides in large amounts by enzymatic degradation of isolated bacterial murein polymer highly specific periplasmic murein-metabolizing enzymes from Escherichia coli are made available. The genes slt, dacB, and mepA, encoding the soluble lytic transglycosylase (Slt), the penicillin-sensitive DD-endopeptidase (PBP4), and the penicillin-insensitive murein endopeptidase A (MepA), were independently fused to the N-terminal encoding sequence of staphylococcal protein A (SpA) under control of the temperature-inducible phage p _R promoter. The SpA fusion proteins were stably over-produced at high levels in E. coli upon temperature induction at 42°C and account for 3% (5 mg SpASlt/l culture), 3% (5 mg SpAPBP4/l culture), and 0.3% (0.5 mg SpAMepA/l culture) of total protein. The SpA fusion proteins, immobilized on IgG Sepharose, are proteolytically sensitive, in vitro, resulting in complete degradation of the SpA portion of the fusion proteins and release of the murein hydrolases in intact and enzymatically active form into the supernatant. Proteolytic degradation could be prevented by p-hydroxymercuribenzoic acid (PHMB) or ethylenediaminetetraacetate (EDTA) suggesting the involvement of the periplasmic protease Pi from E. coli. The immobilized fusion proteins were enzymatically active and could be used for the batch production of biologically active 1,6-anhydro-muropeptides, which were successively separated on HPLC. Isolated murein polymer was degraded quantitatively to monomeric 1,6-anhydro-muropeptides when immunoglobulin G (IgG)-SpASlt was used in combination with IgG-SpAMepA. A combination of IgG-SpASlt with IgG-SpAPBP4 left the 1,6-anhydro-dimers and oligomers being cross-linked via an LD-peptide bond (m-DAP-m-DAP) uncleaved. Correspondence to: W. Keck     

The water relations of Lithobius forficatus and the role of the coxal organs (Myriapoda: Chilopoda)

The water relations of the common brown centipede, Lithobius forficatus (L.), are examined with regard to the possible involvement of the coxal organs in water uptake. Centipedes with blocked and open coxal pores are subjected to a variety of dehydration and rehydration regimes and weight change over time measured. Weight loss and regain is related to body surface area and is not influenced by the coxal organs.
The conclusion of the paper is that the coxal organs of L. forficatus do not contribute significantly to the ability of these animals to recover water lost through cuticular transpiration.     

Occurrence of a Thiothrix sp. Attached to Mayfly Larvae and Presence of Parasitic Bacteria in the Thiothrix sp

Larvae of the mayfly (Drunella grandis [Eaton]) from Diamond Fork Creek, Utah, were covered with a heavy growth of the sulfide-oxidizing bacterium Thiothrix. The bacterium did not seem to harm the mayfly, but the Thiothrix trichomes were parasitized by three morphologically distinct bacteria, two of which were cytoplasmic and one of which was probably periplasmic. At least two of the parasites destroyed the cytoplasmic contents of the Thiothrix sp., thus killing the host cell. Attempts to obtain the parasites in pure culture were unsuccessful.     

Fast-Growing, Aerobic, Heterotrophic Bacteria from the Rhizosphere of Young Sugar Beet Plants

Fast-growing, aerobic, heterotrophic bacteria from the root surface of young sugar beet plants were inventoried. Isolation of the most abundant bacteria from the root surface of each of 1,100 plants between the second and tenth leaf stage yielded 5,600 isolates. These plants originated from different fields in Belgium and Spain. All isolates were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of total cellular proteins. Comparison of protein fingerprints allowed us to inventory the bacteria of individual plants of different fields or leaf stages and to analyze the composition and variability of the rhizobacterial population of young sugar beet plants. Each field harbored a specific population of bacteria which showed a highly hierarchic structure. A small number of bacteria occurring frequently at high densities dominated in each field. The major bacteria were identified as Pseudomonas fluorescens, Xanthomonas maltophilia, Pseudomonas paucimobilis, and Phyllobacterium sp. The former three species showed a high genetic variability as they were represented by different protein fingerprint types on the same or different fields or leaf stages. Twinspan analysis and relative abundance plots showed that the structure and composition of the bacterial populations varied strongly over time. Pseudomonads were typically early colonizers which were later replaced by X. maltophilia or Phyllobacterium sp.